Steroid glucuronides and processes for preparing the same



United States Patent 3,185,682 STEROID GLUCURONIDES AND PROCESSES FOR PREPARING THE SAME Lewis Sarett, Princeton, Robert G. fitrachan, Summit, and Ralph Hirschrnann, Scotch Plains, Nl, assignors to Merck & (10., Inc, Rahway, N.J., a corporation of New Jersey No Drawing. Filed @ct. 4, 1961, Ser. No. 142,792 16 Claims. (Cl. 260-239.55)

This invention isconcerned generallywith novel steroid glucuronides and processes of preparing the same. More particuraly, it relates to glucuronide derivatives of ring A unsaturated l1,l7-bis-oxygenated-20-keto-steroids of the anti-inflammatory pregnane series; with processes of preparing these glucuronide compounds; and with pharmaceutical compositions containing them. These novel ring A unsaturated 11,17-bis-oxygenated -keto-pregnane glucuronides including the preferred ring A unsaturated 11,17-bis-oxygenated ZO-keto-ZI-hydroxy-pregnane 21- glucuronides, while possessing the anti-inflammatory activity characteristic of cortisone, differ from cortisone, hydrocortisone, and their A -derivatives, prednisone and prednisolone, in being remarkably free from the ulcerogenic action, adrenal atrophy, thymus involution and bodyweight-loss side-effects which have resulted from prolonged administration of the aforementioned anti-inflammatory steroids.

The novel 2l-glucuronides of ring A unsaturated 11, 17- bis-oxygenated-ZO-keto-21-hydroxy steroids of the antiinflammatory pregnane series, preferred embodiment of the present invention, may be chemically represented as follows:

wherein ZCH stands for the radical of a ring A unsaturated 11,17-bis-oxygenated 20 keto 21 hydroxysteroid of the pregnane series in which one of the hydrogens at C-21 has been replaced by the glucuronide moiety.

These preferred Zl-glucuronides of ring A unsaturated 1 1,17-bis-oxygenated-20-keto-steroids of the preg nane series may be prepared by reacting the corresponding steroid Zl-free alcohol compound with an alkyl (tri-O- acyl-u-D-glucopyranosyl bromide) -uronate, preferably methyl (tri-O-acetyl-a-D-glucopyranosyl bromide)-uronate, to form the corresponding alkyl (steroid-Zl-yl tri-O-acyl-fl-D-glucopyranosid)-uronate, more particularly the methyl (steroid-Zl-yl tri-O-acetyl-fi-D-glucopy ranosid)-uronate, which is then reacted with an alkaline hydrolyzing agent, thereby forming thecorresponding salt of the steroid 21-glucuronide. a

3,185,682 Patented May 25', 1965 These reactions may be chemically represented as follows:

nane series are conveniently prepared by reacting the steroid 2l-free alcohol and the alkyl (tri-O-acyl-a-D-glucopyranosyl bromide)-uronate together in a hydrocarbon solvent, such as benzene, in the presence of a base such as silver carbonate, and heating the resulting mixture at an elevated temperature. During the course of the reaction, the color of the reaction mixture ordinarily changes from yellow to maroon and, Where silver carbonate is usedas the base, the silver bromide produced during the reaction forms as a suspension in the reaction mixture. The solution containing the steroid-uronate reaction product is then separated from the silver bromide (or other bromide salt by-product's) and the filtered solution is then evaporated to dryness to give the corresponding alkyl (ring A unsaturated 11,l7-bis-oxygenated-ZO-keto-pregname-2 l -yl tri-O-acy1-B-D-glucopyranosid) -uronate, which is conveniently purified by chromatography on activated alumina using mixtures of benzene and chloroform as thesodium met-hoxide, stirring the resultingrnixture togetherat room temperature for a period of about one-half to one hour, and then adding to the reaction solution an aqueous alkaline solution, preferably an aqueous solution of barium hydroxide, and stirring the resulting aqueous mixture at room temperature for an additional one-half to one hour period. When barium hydroxide is used, the barium salt of the ring A unsaturated 11,17-bis-oxygenated-Zl-hydroxy--keto-pregnane 21-glucuronide precipitates from the aqueous mixture and is conveniently recovered by filtration. This barium salt is converted to another pharmacologically acceptable salt, as for example an alkali metal salt such as the sodium salt, an ammonium salt, the choline salt, a basic amino acid salt such as the arginine salt, and the like, by dissolving the barium salt in a minimal amount of water and contacting the aqueous solution with a carboxylic-type cation exchange resin on the appropriate metal cycle, for example the sodium cycle. The resulting aqueous solution of the desired alkali metal salt (e.g. the sodium salt) of the ring A unsaturated 11,17-bis-oxygenated-21- hydroxy-ZO-keto-pregnane 21-glucuronide is then evaporated at low temperature, preferably by freeze-drying, to give the alkali metal salt of the ring A unsaturated 11,17 bis oxygenated-21-hydroxy-20-keto-pregnane 21- glucuronide in substantially pure form.

Alternatively, the hydrolysis reaction is conducted by bringing together, under nitrogen and in solution in an aqueous alcohol, preferably aqueous methanol, the alkyl (ring A unsaturated 11,17-bis-oxy genated-20-keto-pregnane-21-yl tri-O-acyl-fi-D-glucopyranosid)-uronate and a metal alkoxide such as sodium methoxide, stirring the resulting mixture together at room tempearture for a period of about one-half to one hour, during which time the pH changes from above 12 to about 8. It is ordinarily preferred to employ less than the stoichiometric quantity of an alkali metal alkoxide and, particularly, to utilize only one molecular equivalent thereof, whereupon the hydrolysis of the acyl groupings proceeds by ester interchange with the concomitant formation of the methyl acylate which is readily separable from the alkali metal salt of the glucuronide, formed by saponi-iication of the glucuronate ester, by simple evaporation. It is particularly surprising that the reaction proceeds in this way without any appreciable formation of the undesired by-product, the alkali metal'acetate. The reaction mixture is filtered, evaporated to a small volume, diluted with ether, and the precipitate which forms is recovered by filtration, washed with ether, and dried to give the corresponding metal salt (e.g. the sodium salt) of the ring A unsaturated 11,17-bis-oxygenated-21-hyd'roxy-ZO-keto-pregnane 21-glucuronide.

in accordance with these procedures, there are obtained pharmacologically acceptable derivatives, in particular salts with alkali and alkaline earth metals, choline, basic amino acids, and the like, of ring A unsaturated l1,17- bis-oxygenated-Zl-hydroxy-20-keto-pregnane glucuronides as, for example, the glucuronides of the following ring A unsaturated 11,17-bis-oxygenated-2l-hydroxy-ZO-ketosteroids of the anti-inflammatory pregnane series: corti sone, hydrocortisone, and A -isomers thereof; prednisone and prednisolone; A and/or 16-hydroxy derivatives (including acetonides) of any of the foregoing; derivatives of any of these compounds having fluoro, chloro, or bromo substituents attached to the 6, 9, 12 and/or 16- carbon atoms, and/ or methyl substituents attached to the 2, 6, 12, 15 and/or 16 carbon atoms; and the like. Of particular interest are the sodium salts of the 21-.glucuronides of 6,16-dimethyl-cortisone, 6,16-dimethy1-hydrocortisone, and their A derivatives; and [3,2-c1pyrazolo derivatives thereof.

The following examples illustrate methods of carrying out the present invention, but it is to be understood that these examples are given for purposes of illustration and not of limitation. V

Example 1 To a substantially anhydrous mixture of about 0.015 mole of the ring A unsaturated 11,17-bis-oxygenated-20- keto-pregnane 21-free alcohol, approximately 0.03 mole (about 7.40 g.) of silver carbonate and about 350 m1. of benzene, in a distillation equipment provided with an agitator and protected from moisture, is added a substantially anhydrous solution of approximately 0.02 mole (about 8.00 g.) of methyl (tri-O-acetyl-wD-glucopyranosyl bromide)-uronate in 50 ml. of benzene over a period of one hour while distilling benzene from the reaction mixture at a substantially equal rate. During the course of the reaction, the color of the mixture changes from yellow to maroon. The distillation of the benzene is continued for an additional forty minute period, the reaction mixture is cooled to room temperature, and the silver bromide formed during the reaction is separated from the mixture by filtration. The filtered solution is evaporated to dryness in vacuo to give the corresponding methyl (ring A unsaturated 1 1,17-bis-oxygenated-ZO-keto-pregnane-2l-yl tri-O-acetyl-fi-D-glucopyranosid)-uronate in a yield approaching that theoretically obtainable.

This material is conveniently purified by dissolving it in benzene-chloroform (9:1), passing the resulting solution through a column containing approximately 350 g. of neutral activated alumina, and eluting the resulting adsorbate, first with 800 ml. of benzene-chloroform (9:1), then with 800 ml. of benzene-chloroform (4:1) (which solvents do not remove appreciable steroidal product from the column) and finally with 4.5 liters of benzene-chloroform (7:3). Evaporation of the latter eluate gives the methyl (ring A unsaturated 11,17-bisoxygenated 20 keto-pregnane 21 yl-tri-O-acetyl-[S-D- glucopyranosid)-uronate in substantially pure form in a yield of over 50% of theory.

In accordance with the above procedure, but using 0.02 mole of ethyl (tri-O-acetyl-a-D-glucopyranosyl bromide)- uronate instead of methyl (tri-O-acetyl-a-D-glucopyranosyl bromide)-uronate, there is obtained the corresponding ethyl (ring A unsaturated 11,17-bis-oxygenated-20- keto-pregnane 21 yl tri-O-acetyl-fi-D-glucopyranosid)- uronate.

Similarly, using 0.02 mole of another alkyl (tri-O-alkanoyl-a-D-glucopyranosyl bromide)-uronate, as for example butyl (tri-O-propionyl-iz-D-glucopyranosyl bromide)-uronate in the foregoing reaction, there is obtained the corresponding alkyl (ring A unsturated 11,17-bis-oxygenated-ZO-keto-pregnane-Z1-yl tri-O-alkanoyl-B-D-glucopyranosid)-uronate, as for example butyl (ring A unsaturated 11,l7-bis-oxygenated-ZO-keto-pregnane-2l-yl tri-O- propionyl-fi-D-glucopyranosid)-uronate. Where the ring A unsaturated 11,17-bis-oxygenated-ZO-keto-pregnane 21- ree alcohol, reacted with silver carbonate and methyl (tri-O-acetyl-zx-D-glucopyranosyl bromide)-uronate in accordance with the foregoing procedure, is hydrocortisone, the product obtained is methyl (11fl,17cadll1Yd10XY-4- pregnene-3,20-dione-2l-yl tri-O-acetyl-B-D-glucopyr-anosid)-uronate. Similarly, when prednisolone is used as the ring A unsaturated 11,17-bis-oxygenated-ZO-keto-pregnane-2l-free alcohol, there is obtained methyl (115,170:- dihydroxy-1,4-pregnadiene-3,20-dione-21-yl tri-O-acetylfi-D-glucopyranosid)-uronate; a 243 m E percent 197. Similarly, when loot-methyl hydrocortisone is used as the ring A unsaturated 11,l7-bis-oxygenated-20-ketopregnane ZI-free alcohol, there is obtained methyl (16w methyl-1l5,17a-dihydroxy 4 pregnene-3,20-dione-2l-yl tri-O-acetyl-h-D-glucopyranosid)-uronate; k 240 m E percent approximately 200.

Utilizing the foregoing procedure, and as reactants, the particular ring A unsaturated 11,17-bis-oxygenated-20- keto-pregnane 21-free alcohol and alkyl (tri-O-acyLa-l glucopyranosyl bromide)-uronate indicated in columns 2 and 3 of the following table, there is obtained the corre sponding alkyl (ring A unsaturated 11,17-bis-oxygenated- Li a ZO-keto-pregnane 21 yl tri-O-acyl-fi-D-glucopyranosid)- uronate shown in column 4 thereof:

6 ring A unsaturated 1 1,l7-bis-oxygenated-2l-hydroxy-20= kcto-pregnane-Zl-glucuronide is dissolved in a minimum Ring A unsaturated 11,17-bis- Alkyl (tri-O-acyLa-D-glucopy- Alkyl (Ring .A. unsaturated 11,17-bis- Expt. N0, oxygenated 20-keto-pregnane 21- rauosyl bromide)-uronate oxygenated-20-keto-preguane 21-yl trifree alcohol O-aeyl-B-D-glucopyranosld)-uronate 1 lfia-methyl prednisolone Methyl (trl-O-acetyl-oi-D-glueo Methyl (lfia-methyl-lla, l7a-dihydr0xy-1,

pyransy1bromide)-uronate. 4-pregnadiene-3,20-dione-2l-yl tri-O-acetyl-B-D-glucopyran0sid)-uronate.

2 IGfl-methyl prednisolone. Methyl (tn-O-acetyl-a-D-gluco- Methyl (IGB-methyl-llfi, 17a-dihydroxy-1,+

pyranosyl bromide)-uronate. pregnadiene-3,20-dione-21-yl tri-O-aeetyl- V fl-D-glucopyranosid)-uronate. V

pyranosyl bronudeJ-uronate. droxy-l,4-pregnadiene-3,20-dione-21-yl tri- O-acetyl-fl-D-gluc0pyranosid)-ur0nate.

pyrauosyl hro1nlde)-uronate. droxy-l ,4-pregnadiene-3.20-dioue-2l-yl tri- O-aeetyl-B-D-glucopyranosid)-uronate.

5 Got-methyl prednisolone Ethyl (tn-O-prop1onyl-c -D-gluc0- Ethyl (fiwmethyl-llfid7wdihydroxy-1A- pyranosyl bromide)-uronate. pregnadiene)-3,20-dion-2l-yl tri-O-propionyl-fl-D-g1ye0pyran0sid)-ur0nate.

6 fia-fluoro pr nisolon Ethyl (tri-O-propionyl-a-D-gluco- Ethyl (swallo -11g, mnm q,

pyranosyl bromide)-uronate. pregnadieue-3,20-dione-2l-yl tri-O-propionylfl-D-glucopyranosid)-u.ronate.

7 6a, isa-dimethyl prediusolonenm Methyl (tn-O-acetyl-a-D-zlu- Methyl (6oz. lea-diniethyldle, I'Ta-dlhycopyranosyl bromide)-ur0nate. droxy-l, 4-pregnadiene-3, 20-dione-2l-yl trlO-acetyl-B-D-g1ueopyranosid)- 4 uronate,

3 6, lfia-dimethyl-llfl, 170., 21- Methyl (trnO-acetyl-a-D-glu- Methyl (6, lfia-dimethyl-llfi, 17a-dihytrihydroxy-l, 4, fi-pregnatrienecopyranosyl bromide)-uronate. droxy-l, 4, 6-pregnatriene-3, QO-dione- 3, zddione, 21-yl ttri-O-aeetyl-B-D-glueopyranosid)- urona e.

9 9a-fiuoro-lfia-hydroxy-predniso- Methyl (tri-O-acetyl-a-D-glu- Methyl(9a-fluoro-11,6, 161x, 17a-trihydroxy-1,

lone. eopyranosyl bromide)-uronate. -pregnadiene-Z, 20-di0ne 2l-yl tri-O- aeetyl-dD-glucopyran0sid)-urauate.

10 Qa-fiuotO-fi, lfia-dl'fnGthyl-IIB, Butyl (tri-O-benzoyl-a-D-glw Butyl (9a-fluoro-6, IGcr-dll'l'lthYl-IIB, 17a- 17a, Zl-trihydroxy-l, 4, G-pregcopyrauosyl bromide)-urouate. dihydroxy-l, 4, 6-pregnatriene-3, 20- natriene-B, Z-D-dione. dione-Zl-yl tri-O-benzoy1-fl-D-gluc0- pyran0sid)-uronate.

11 17a, 21-dihydroxy-l-allopregene- Propyl (tri-O-butyryl-a-D-glu- Propyl (l7a-hydroxy-1-allopregnene-3, 11,

3, ll, ZO-trione. copyranesyl bromide)-uronate ZO-trlone-Zl-yl tri-O-butyryhB-D- glucopyranosid)-uronate.

12 ga-fluoro-llfi, 17a, ZI-trihydroxy- Methyl (tri-O-acetyl-a-D-glu- Methyl (9a-fiuor0-11fl, l7a-dihydroxy-l- 1-pregnene-3, 20-dione. eopyranosyl bromide) uronate. pregnene3, 20-dione-2l,y1 tri-O-acetyl- B-D-glucopyrauosid)-uronate.

13 115, 17a, 21-tril1ydr0xy' [3, 2-0] Methyl (tri-O-aeetyl-a-D-glu- Methyl (11B. l7a-dihydr0xy- I3, 2-e] pyrazpyrazolo-tpregnene-ZO-one. copyranosyl bromide)-ur0nate. ole-443mgnene-20 0ne-21-yl tri-O- 14 g nner -115, 17a, ZI-trihydroxy- Methyl (tri-O-acetyl-a-D-gluco- Methyl (9a-fiuoro-ll5, l7a-dihydroxy-[3,2-

[3,2-e] pyrazolo-tpregnenewpyranosyl bromide)-uronate. e] pyrazolo-4-pregnene-20-one-2l-yl trione. O-acetyl-B-D-glucopyran0sid)-uronate.

15 9a fluoro 16i1-methyl 11B, 170:, 21- Methyl (tn-O-ecetyl-a-D-gluco- Methyl (9a-fiuoro-lSa-methyl-llfi,17a-

trihydroxy-[3,2-c] pyrazolopyranosyl bromide)-urenate. dihydroxy-[3,2c] pyrazolokpregnenepregnene-'0ne. 20-one-2l-yl tri-O-acety1--D-glucopyranosid)-uronate.

15 lfi -methyl-llfl, 170i, 21-trihydroxy- Methyl (tri-O-aeetyl-a-D-glueo- Methyl (lfia-methyl-llfi, 17a-dihydroxy [3,2-0] pyraz0l0-4-pregnene20- pyranosyl bromide)-uronate. [3,2 0] pyrazolo-ipregncnea0-one'21-y1 one. tri-O-aCetyl-B-D-glucopyran0sid)- uronate.

17 c ti Methyl (tri-O-acetyl-a-D--gluco Methyl (17a-hydroxy-4-pregnene-3,l1,20-

pyranosyl hromide)-ur0nate. trione-Zl-yl tri-O-aeetyl-fl-D-glueopyran- W osid)-uronate.

pyranosid)-uronate.

19 6,lfia-dimethyl-llfl,1701,21-tll- Methyl (trl-O-aeetyl-a-D- lu-co Methyl (6,lfia-dirnethyl-llfl,l'ia-dihydroxyhydroxy-2-phenyl-[3,2-e] pyrazpyranosyl bromide) -u.ronate. 2-phenyl-[3-2 c] pyrazolo-4,6-pregna- 010-4,itpregnediene-iil-one diene-20-one-2l-yl triO-acetyl-B-D-glueopyranosid)-uronate.

2g 6,16a-di111ethYI-l1fi,17a,21-t1lhy- Methly (tri-O-aeetyl-a-D-gluco- Methyl (6,16a-dimethyl-llfl,l7a dihydroxydroxy-4,6-pregnadiene-3,20- pyranosyl bromide-uronate. 4,G-pregnadlene-3,20-dione-2l-yl tri-O- dione. acetyl-fl-D-gluc0pyranosid)-uronate.

Example 2 Approximately 0.005 mole of alkyl (ring A unsaturated l 1,17 -bisoxygenated-ZO-keto-pregnante-2l-yl tri-O-acyl-fl- D-glucopyranosid)-uronate is dissolved in about 70 ml. of methanol, and to this solution is added about 20 ml. of an approximately 1 N solution of sodium rnethoxide in methanol. The resulting mixture is stirred under nitrogen at room temperature for approximately minutes. To the resulting mixture is then added, drop-Wise with stirring, about 250 ml. of an approximately 0.1 N aqueous solution of barium hydroxide, whereupon a precipitate separates. The resulting slurry is stirred at room temperature for an additional period or about 30 minutes, and the precipitated material is recoveredby filtration, Washed with methanol until neutral, and dried to give approximately 0.60l'mole of the barium salt of the ring A unsaturated ll,l7-bis-oxygenated 21 hydroxy-ZO-keto-pregnane-2l glucuronide. by cooling the mother liquor and allowing it to stand at G C.

Approximately 0.001 mole of the barium salt of the Additional material is recovered quantity of Water at room temperature, and the resulting solution is filtered. To this filtered solution is added about 30 g. of a carboxylic type cation exchange resin (eg. Amberlite IRC-SO resin manufactured by Rohm & Haas Company, Philadelphia, Pennsylvania), on the sodium cycle, and the mixture is stirred at room temperature for about thirty minutes. The reaction solution is separated from the resin (as by filtration or decantation), extracted with a halogenated hydrocarbon solvent such as methylene chloride (thereby extracting ring A unsaturated l1,17-bis-oxygenated-2l-l1ydroxy-20-keto-pregnane which may be present), and evaporated to dryness at low temperature (e.g. by freeze-drying) to give approximately 0.001 mole of the sodium salt of the ring A unsaturated ll,l7-bis-oxygenated-2l-hydroXy-ZO-keto pregnane 21- glucuronide.

In accordance with the above procedure, but using the carboxylic cation exchange resin on another metal cation cycle, for example an alkali metal cation such as potassium (14+), an alkaline earth metal cation such as talciuni (Ca++), and the like (instead of the sodium cycle) there is obtained the corresponding metal salt of the ring A unsaturated-ll, l7-bis oXygenated-2l-hydroxy 20 ketopregnane-Zl-glucuronide, such as the potassium salt of the ring A unsaturated-11,17-bis-oxygenated-2l-hydroxy 20- keto-pregnane-2l-glucuronide, the calcium salt of the ring A unsaturated-1 l, l 7-bis-oxygenated-2 l-hydroxy-ZO-ketopregnane-Zl-glucuronide, and the like.

In accordance with the above procedure, but using methyl (ring A unsaturated-l1,l7-bis-oXygenated-20-keto pregnane-21-yl tri-O-acetyl-fi-D-glucopyranosid)-uronate as starting material and the carboXylic cation exchange resin on the sodium cycle, there is obtained the sodium salt of the corresponding ring A unsaturated-11,17-bisoxygenated-21-hydroXy-2O-keto-pregnane-Z l-glucuronide.

Where the starting material utilized in the foregoing procedure is methyl (l1p,l7oa-dihydroXy-4-pregnene3,20- dione-Zl-yl tri-O-acetyl-5-D-glucopyranosid)-uronate and the carboxylic cation exchange resin is used on the sodium cycle, the product obtained is the sodium salt of 1 1B,170:,21-trihydroxy-4-pregnene-3,ZO-dione 21-glucuronide. Similarly, when the alkyl (ring A unsaturated-ll, 17-bis-oxygenated-ZO-keto-pregnane-Zl-yl tri-O-acyl-[i D- glucopyranosid)-uronate used as starting material in the foregoing procedure is methyl (ll,B,17a-dihydroxyl 1,4- pregnadiene-3,20-dione-2l-yl tri-O-acetyl-[i-D glucopyranosid)-uronate and the carboxylic cation exchange resin is used on the sodium cycle, there is obtained the sodium salt of 11,8,17m,2l-trihydroxy-1,4-pregnadiene-3,ZO-dione ZI-glucuronide A 246 mu; E percent 167. Similarly, when methyl (la-methyl-l lfi,17ot-dihydroxy-4-pregnene- 3,20-dione-2l-yl tri-Q-acetyl-fi-E-glucopyranosid) -uronate is used as starting material and the carhoxylic cation exchange resin is used on the sodium cycle, there is obtained the sodium salt of lowmethyl-llfi,l7a,2l-trihydroxy-4-pregnene-3,ZO-dione 2 l-glucuronide.

Utilizing the foregoing procedure, and as starting material, the particular alkyl (ring A unsaturated-11,17-bisoxygenated-20-keto-pregnane-2i-yl tri-O-acyl-fl-D glu copyranosid)-uronate indicated in column 2 of the following table, there is obtained the sodium salt of the corresponding ring A unsaturated-11,17-bis-oxygenated 21- hydroxy-ZO-keto-pregnane Zl-glucuronide shown in column 3:

Alkyl (Ring A unsaturated-11,17- bisoxygenated-QU-keto pregnane 21-yl tri-O-acyl--D-glucopyranosid)-uronate Ring A unsaturated-11,17-his-oxygenated 21 hydroxy 2O ketopregnane 2l-glucuronide sodium salt lfia-rnethyl-llfi, 17a, 21-trihydroXy-1A- prcgnadiene 3,20 dione 21 glueur onide sodium salt.

16,6 methyl 115, 17a, 21 trihydroxy 1,4 pregnadiene 3,20 dione 21 glucuronide sodium salt.

9a-fi u0r0 16a-methyl-11/3, 17a, 21- trihydroxy-1,4-pregnadiene-3,2- tione 2l-glucuronide sodium salt.

Qa-iiuoro-lGfi-mcthyl-llfi, 17m, 21-

tr hydroxyd,4-pregnadiene-3,20- drone 21-glucuronide sodium salt.

(la-methyl-llfl, l7a,2l-trihydroxy-l, 4-pregnadien e-3,20dione 2l-glucuronide sodium salt.

(SaJGa-dimethyI-Hfl, 170:, 21-trihydroxy-l,4-pregnadiene-3,20dione 2l-glucuronide sodium salt.

6,l6a-dimethyll1 9,17a 21-trihydr0xyl,4,(i-pregfnatnene-8QO-dione 2lglucuronide sodium salt.

9a fl uoro-S,loa-dimethyliifl,1704,21-

trihydroxy-l,4,6-pregnatriene-3.20- dione 2l-glucuronidc sodium salt.

17a,21- dihydroxy-l-allopregncire-3,11.

20-tr1o11e Zl-glucuronidc sodium salt.

Qa-fluoro-l1,6,17a,2l-trihydroxy-[32-e] pyrazolo--pregneno-ZO-one 2l-glucuronide sodium salt.

1701,21-dihydroxy-4-preg11ene-3,11,20- tnone-Zl-glueuronide sodium salt.

one 2l-glucur0nide sodium salt.

Allryl (Ring A unsaturated-l1,l7- Ring A unsaturated-l1,17-bis-oxygon- Expt. bis oxygenatedketopregnaneated 21 hydroxy 20 keto preg- N 0. 21-yl tri-O-acyl-B-D-gluc0pyran0- nane 21-g1ucuronide sodium salt sid)-uronate l9 Methyl (6 .16 dimethyl-l1B',17a-dihy- 6,16a-dimethyl-llfi,17a,2l-trihydroxydroxy-2 -phenyl [3,2-c] pyrazo1o-4.6- 2-phenyl-[3,2-c] pyrazo1o-4,6-pregnapregnadine-ZO-one-Zl-yl tri-O-aeetyldiene-ZO-one 21-g1ucurouide sodium ,o-D-glueopyranosid)-uronate. s 20 Methyl-(6,lfia-dimethyl-l1f3,17a-dihy- 6,lda-dimethyl-l1B,l7a,2l-trihydroxydroxy-4,6-pregnadine-3,20 dione-2l-. 4,6-pregnadiene-3,20-dione Zl-gluyl tri-1?-acetyl-B D-glueopyran0sid)- curonide sodium salt.

uroua e.

Exmnple 3 pproximately 0.006 mole of alkyl (ring A unsaturated 1l,17-bisoxygenated ZO-keto-pregnane-Z1-yl tri-O-acyl 4igiucopyranosid)-uronate is dissolved in about 40 ml. of methanol, and to this solution is added about 1.5 ml. water and about 2.5 ml. of an approximately 2.4 N methanolic sodium methoxide solution. reaction solution immediately changes from yellow to orange. The reaction solution is then stirred at room temperature under a nitrogen atmosphere for a period of approximately minutes. The pH of the solution changes from above 12 to approximately 8. The reaction mixture is filtered, evaporated in vacuo to slightly less than /2 the original volume (about 20 ml). Diethyl other is added, whereupon a precipitate forms, and the addition of the ether is continued until no further precipitation occurs. The precipitated material is recovered, washed with ether, and dried to give the sodium salt of the ring A unsaturated l1,17-bis-oxygenated-21-hydroxy ZO-izeto-pregnane Zl-glucuronide in a yield of approximately 80% of-that theoretically obtainable.

in accordance with the above procedure, but using an aqueous methanolic solution containing another alkali metal alkoxide, such as potassium ethoxide (instead of the sodium methoxide), there is obtained the corresponding alkali metal salt of the ring A unsaturated 11,17-bisoxygenated-21-hydroxy-20-keto-pregnane 21- glucuronide, such as the potassium salt of the ring A unsaturatedl 1,17- bis-oxygenated-21-hydroxy-20-kcto-pregnane ZI-glucuronide, and the like.

In accordance with the above procedure but using methyl (ring A unsaturated 11,17-bis-oxygenated-ZO-ketopregnane-Z l-yl tri-O-acetylfi-D-glucopyranosid -uronate as the alkyl (ring A unsaturated 11,1'7-bis-oxygenated-20- keto-pregnane-2l-yl tri-O-acyl=fi-D-glucopyranosid)-uronate and a methanolic solution of sodium methoxide, as the reactants, there is obtained the sodium salt of the corresponding ring A unsaturated 11,17-bis-oxygenated- 21-hydroxy-20-keto-pregn ane 2'i-glucuronide.

Where the starting material reactedwith the aqueous methanolic solution of sodium methoxide in the fore going procedure is methyl (.1lfl,17oa dihydroxy-4-pregnene 3,20 dione 21 yl tri O acetyl ,8 D glucopyranosid)-uronate, the product obtained is the sodium salt of 1l,8,l7ot,21-trihydroxy-4-pregnene-3,ZO-dione 21- glucuronide. Similarly, when the alkyl (ring A unsaturated 1 1,l7 bis oxygenated 20 keto pregnane 21- yl tri-0-acyl-/3-D-glucopyranosid)uronate, used as starting material in the foregoing reaction with sodium methoxide in aqueous methanol, is methyl (11,8,17a-dihydroxy 1,4 pregnadiene 3,20 dione 21 yl tri- O-acetyi-B-D-glucopyranosid)-uronate, there is obtained the sodium salt of 115,17a,21-trihydroxy'1,4-pregna-' dione-3,20-dione 21-glucuronide. Similarly, when methyl (160a methyl 116,170 dihydroxy 4 pregnene 3,20- dione 21 yl tri O acetyl [3 D glucopyranosid)- uronate is used as starting material in the foregoing reaction with sodium methoxide in aqueous methanol, there is obtained the sodium salt of 16tx-methyl-11/3,17u, 21-trihydroxy-4-pregnene-3,20-dione Zl-glucuronide.

The alkyl (tri-O-acylx-D-glucopyranosyl bromide)- uronates such as methyl (tri-O-acetyl-a-Dglucopyranosyl The color of the bromide)-uronate, used as starting materials in the foregoing examples are prepared as follows: a mixture of about 18 g. of glucuronolactone, 0.15 g. of sodium methoxide and ml. of methanol is stirred at room temperature for a period of approximately 1 hour. The resulting reaction solution is evaporated to a vast syrup in vacuo. This syrup is then dissolved in approximately 70 ml. of acetic anhydride, and to this solution is added, dropwise with stirring, a solution of 0.3 ml. of perchloric acid in 10 ml. of acetic anhydride, while maintaining the reaction temperature below about 40 C. The resulting mixture is allowed to stand at room temperature for a period of about 15 hours, an additional 0.1 ml. of perchloric acid is added, and the resulting solution cooled to about 0 C. and maintained at that temperature for an additional 15-hour period. The crystalline material which separates is recovered by filtration, washed with ether, and recrystallized from ethanol to give substantially pure methyl tetra-O-acetyl-[3-D-glucopyranuronate. Utilizing the foregoing procedure but using (1) in place of the 100 ml. of methanol there employed, approximately 100ml. of another alcohol such. as ethanol, propanol, butanol, benzyl alcohol and the like, (2) in place of the 0.15 g; of sodium methoxide, about 0.l50.2 g. of a sodium alkoxide corresponding to the alcohol utilized, i.e. sodium ethoxide, sodium propoxide, sodium butoxide, sodium benzoxide, respectively, and (3) in place of the 70 ml. of acetic anhydride, about 70-100 ml. of another acyl anhydride such as propionic anhydride, butyric anhydride, and the like, there is obtained the corresponding alkyl tetra-O-acyl-,8-D-glucopyranuronate such as ethyl tetra-O-propionyl-fl-D-glucopyranuronate; butyl tetra O butyryl 9 D glucopyranuronate; benzyl tetra-O-acetyl-fi-D-glucopyranuronate, and the like.

A solution of about 50 g. of methyl tetra-O-acetyl-B- D-glucopyranuronate in about 200 ml. of 30% hydrobromic acid in acetic acid is cooled to about 0 C. and allowed to stand at that temperature for a period of about 15 hours. The solvent is evaporated from the reaction solution in vacuo at a temperature of below about 40 C., and the residual material is dissolved in 100 ml. of chloroform. The chloroform solution is washed with cold saturated sodium bicarbonate solution, then with water, and dried over anhydrous sodium sulfate. The washed dried chloroform solution is evaporated in vacuo to a syrup. The residual syrup is dissolved in about ml. of absolute ethanol, treated with activated charcoal, filtered, and the filtered solution cooled to about 0 C. and allowed to stand atthat temperature for about '15 hours. The crystalline material which separates is recovered by filtration and dried to give substantially pure methyl (tri-O-acetyl-ot-D-g'lucopyranosyl bromide)-uronate.

Similarly, utilizing other alkyl tetra-O-acyl-fi-D-glucopyranuronates such as ethyl tetra-O-propionyl-B-D-glucopyranuronate; butyl tetra O butyryl B D glucopyranuronate; benzyl tetra O acetyl [3 D glucopyranuronate, and the like in the foregoing reaction with the 30% hydrobromic acid in acetic acid, there is obtained the corresponding alkyl (tri-O-acyl-a-D-glueopy ranosyl bromide)-uronate such as ethyl (tri-O-propionyla-D-glucopyranosyl bromide)-uronate; butyl' (tri-O-bui i. tyryl-a-D-glucopyranosyl bromide)-uronate; benzyl (tri- O-acetyl-a-D-glucopyranosyl bromide)-uronate, and the like.

These new ring A unsaturated 11,17-bis-oxygenated 20- keto-steroid glucoronides of the anti-infiammatory pregnane series possess anti-inflammatory activity characteristic of cortisone, and their use results in little salt and water retention or adrenal atrophy. They are, in addition, remarkably free from thymus involution and bodyweight-loss side effects ordinarily accompanying the prolonged administration of anti-inflammatory steroids. They are normally administered in a daily maintenance dosage range comparable with that utilized in the administration of the corresponding ring A unsaturated 11,17-bisoxygenated 20-keto steroid, e.g. from about 25 to 75 mg. per day for cortisone glucoronides or hydrocortisone glucuronides; about 2.5 to mg. per day for prednisolone glucuronide; and about 0.25 to 5 mg. per day in the case of dexamethasone glucuronide, although, in view of their selective anti-inflammatory action (substantially unaccompanied by undesired side effects), they may, in the case of aggravated inflammatory conditions, be administered in substantially higher dosages Without attendant risk of side effects; and, in milder conditions, may often be administered in substantially lower dosages in View of their pronounced anti-inflammatory action directly at the site of the inflammation. While, due to their relative instability in acid solution, these ring A unsaturated 11,17- bis-oxygenated -keto-steroid glucuronides are ordinarily administered by injection, they may also be administered by the oral route, preferably admixed with, and/ or coated by, an alkaline-reacting pharmacologically-acceptable buffer substance, or in an enteric-coated capsule.

, The ring A unsaturated 1l,17-bis-oxygenated-l6-methyl-20-ketopregnane 21-free alcohols, more particularly the 1601- and 16,8-methyl derivatives of cortisone and hydrocortisone, their A derivatives, 6-methyl and 6-methyl- A derivatives of the foregoing, and [3,2-c] pyrazoleanalogues thereof, used as starting materials in the foregoing examples, are prepared in accordance with the following procedures:

A solution of 10.22 g. of methyl iodide in 50 ml. of ether is added to 1.73 g. of magnesium in 50- ml. of ether. To the resulting ethereal solution of methyl magnesium iodide, maintained under a nitrogen atmosphere, is added 0.045 g. of anhydrous cuprous chloride. To this mixture is added, over a period of about one hour, during which,

period the reaction mixture is stirred vigorously and maintained at approximately room temperature, a solution of about 5.6 g. of 3ct-acetoxy-16-pregnene-11,29-dione in 175 ml. of ether. A white granular solid separates during this addition. The resulting mixture is heated under gentle reflux for two hours after which the reaction mixture is cooled, and 125 ml. of saturated, aqueous ammonium chloride solution is added followed by 200 ml. of ether. The layers are separated, and the ethereal layer is washed with three 50 ml. portions of water. The washed ethereal layer is dried, and the solvent evaporated in vacuo to give a brown viscous oil. The latter material is heated for 15 minutes at 6070 C. with a mixture of 25 ml. of acetic anhydride and 25 ml. pyridine and the a-cetylated product is purified by chromatography on acidwashed alumina followed by crystallization from petroleum ether to give approximately 1.5 g. of substantially pure 3ot-acetoxyl6m-methyl-pregnane-11,20-dione.

To a solution of 0.8 g. of 3et-acetoxy-l6a-methyl-pregnane-11,20-dione in ml. of methanol is added 1.5 ml. of concentrated aqueous hydrochloric acid and the resulting solution is stirred overnight at about 25 C. The reaction solution is evaporatedin vacuo at 25 C. to a small volume, and the concentrated solution is poured into ml. of ice water. The white solid which precipitates is recovered by filtration, washed with water and recrystallized from ethyl acetate to give 3a-hydr0xy-l6u-methylpregnane-l 1,20-dione.

A solution or" 22 g. of 3ot-acetoxy-16ot-methyl-pregnane- 11,20-dione and 1 g. of p-toluene-sulfonic acid in 250 ml. of acetic anhydride is heated at reflux under nitrogen for a period of approximately 3 days. Two grams of potassium acetate (anhydrous) is added, and the volatile solvents are separated by distillation in vacuo. The residual material is extracted with benzene, and the benzene extract is filtered to remove insoluble material. The benzene extracts are evaporated to a volume of 100 ml. and petroleum ether is added to the cloud point. The resulting solution is adsorbed on 660 g. of acid-washed alumina; the alumina adsorbate is then washed with 2 liters of petroleum ether. The adsorbate is then eluted with 15 petroleum ether-ether mixture, and the first four liters of eluate is collected, and evaporated to dryness in vacuo to give a mixture of enol acetates containing 3a,20-diacetoxy-16otmethyl-17 (20) -pregnene-i 1- one. This mixture of enolates, weighing approximately 14 g., is dissolved in 50 ml. of benzene and treated with an excess of per-benzoic acid over a 16-hour period. The reaction mixture is shaken with dilute aqueous potassium hydroxide solution until the benzene layer is free of per-benzoic acid; the benzene layer is then washed with water until neutral, dried, and the solvent evaporated in vacuo to give a crystalline material, 17a,20-cpoxy- 3u,20-diacetoxy1u-methyl-pregnane-ll-one. The latter material is dissolved, Without purification, in 200 ml. of methanol, 120 ml. of water and 10 g. of potassium bicarbonate, and the resulting solution is heated at reflux under nitrogen for a period of 16 hours. The methanol is evaporated from the hydrolysis solution in vacuo, and the residual oil is extracted from the resulting aqueous solution with chloroform. The chloroform extract is washed with water to neutrality, dried, and the chloroform is evaporated under reduced pressure. The residual oil is triturated with ether, and the crystalline material thus formed is recrystallized from ethyl acetate-petroleum ether to give 3a,17u-dihydroxy-16a-methyl-pregnane- 11,20-dione.

To a solution of 7.0 g. of 3a,17a-dihydroxy-Mat-methylpregnane-11,20-dione in 50 ml. of chloroform is added dropwise with stirring a solution containing 3.36 g. of bromine in 24.2 ml. of chloroform over a period of about 60 minutes. The reaction mixture is dissolved in 200 ml. of ethyl acetate, and the resulting solution washed with water until neutral, dried, and the solvents evaporated therefrom in vacuo. The residual crude material is dissolved in a minimum quantity of ethyl acetate, the resulting solution is diluted with ether, and the mixture is stirred until crystals form. The crystalline product is recovered by filtration and washed, by slurrying, with 50:50 ether-petroleum ether mixture to give about 5 g. of 21-bromo-3a,17m dihydroxy 16oz methyl-pregnanc- 11,20-dione.

This 5 g. of 21-bromo-3u,17u-dihydroxy-Mot-methylpregnane-lLZO-dione is mixed with 5.0 g. of anhydrous potassium acetate, 4.0 g. of sodium iodide and 0.03 ml. of glacial acetic acid, and ml. of acetone is added to the resulting mixture. This mixture is then heated at reflux, with stirring, for a period of about 16 hours, and the reactionmixture is cooled, filtered, and the insoluble material is washed with acetone. The filtered solution is evaporated in vacuo thereby removing the solvents, and the residual material is slurried with water, and the aqueous mixture extracted with ethyl acetate. The ethyl acetate extract is Washed with water to neutrality, dried, and the solvent is evaporated in vacuo to give an oil. This oil is crystallized from ether, and recrystallized from ethyl acetate-ether to give 3a,17a,21-trihydroxy-I6a-methyl-pregnane-11,20-dione 21-acetate.

A solution of 400 mg. of 3u,17ct,2l-trihydroxy-16arnethyl-pregnane-l1,20-dione 21-acetate in 4 ml. of pyridine is added to the complex formed by the addition of 400 mg. of chromium trioxide to 4 ml. of pyridine. The

mixture is swirled until thoroughly mixed, and then al lowed to stand at room temperature overnight. The reaction mixture is poured into water, and the aqueous mixture is extracted with ether, and then twice with ethyl acetate. The combined ether and ethyl acetate extracts are washed with dilute aqueous sulfuric acid at about C., and then with water until neutral. The organic solvent layer is then dried, the solvents are evaporated therefromin vacuo, and the residual crystalline material is purified. by crystallization from ethyl acet-ate to give 17a,2-1- dihydroxy 16a methyl-pregnane- 3,11,20-trione 21.-acetate.

To 100 mg. of 17a,2l-dihydroxy-l6oc-methyl-pregnane- 3,11,20-trione 21-acetate dissolved in 2 ml. of chloroform and; 2.25 ml. of glacial acetic acid, at a temperature of 55 C., is added: two drops of a 0.001 N solution of dry HBr in. glacial acetic acid. To about 0.38 ml. of 0.001 N HBr in glacial acetic acid, at -55 C., is added 043 ml. of a solution containing 40 mg, of bromine in chloroform, and the resulting solution is added, over about a 10-minute period, to the solution ot'the steroid, While maintaining the reaction mixture at about 55 C. The reaction mixture is allowed to stand at 5,5 C. for about one-half hour; a solution containing 250 mg. of sodium. acetate; in 3 ml. of water is added, and the resulting mixture is stirred for about 5 minutes. Five milliliters of water are then added, and the aqueous mixture is extractedwith ethyl acetate. The. ethyl acetate extractis washed with aqueous sodium bicarbonate solution to neutrality, then with water, dried, and the solvent is evaporated in vacuo. The residual material is dissolved in. 2 ml. of acetone, and to the solution is added 25 mg. of sodium bromide and 1 ml. of water. The resulting mixture is heated under reflux for a period of, about 5 hours, the reaction mixture is cooled, and the acetone is evaporated in vacuo. The residual material is extracted into ether, the ether'extract is washed with, water, dried, and the solvent is evaporated to a volume of about 1 ml.; petroleum ether is added to this solution, and the crystalline material which separates is recovered and dried to give approximately 90 mg. of 4- bromo-17a,2l-dihydroxy-16a methyl-pregnane-3,11,20- trione 21-acetate. v

A mixture of 48 mg. of semicarbazide, 48 mg. of 4- bromo-17u,21 dihydroxy 16 umethyl-pregnane3,l1,20- trione 21-acetate and 0.6 ml. of ethanol is heated under reflux in contact with a nitrogen atmosphere for a period of about three days, and the reaction solution is evaporated to a small volume, diluted with water and the crystallinematerial recovered and purified by recrystallization from aqueous methanol to give 3,20-bis-semicaroa- Zi-Cl0l7o,21 dihydroxy 16a methyl-4-pregnene-3,l1,20- trione 2l-acetate. Fifty milligrams of 3,20 -bis-semicarbazidol 7oc,2l dihydroxy-1 6a-inethyl-4-pregnene-3 ,1 1,20- trione 21-acetate is dissolved in a mixture of 1.0 cc. of benzene and 1.0 cc. of 1.1 N methanolic potassium hydioxide, and the solution is allowed to stand at room temperature for a period of about 10 minutes. The solution is then acidified with acetic acid, the benzene is evaporated in vacuo,, and the residual material is recrystallized from ethyl acetate to give 3,,20 bis-sernicarbazido-17a,21-dihydroxy-l6otmethyl-4-pregnene-3,1 1,20-trione.

A mixture of 60 mg. of 3,20-bis-semicarbazido-17a,21- dihyd'roxy-l6a-rnethyl-4-pregnene-3J1,20-trione, 0.2 ml. of dimethylformamide, 0.6 ml. of chloroform, and 1.5 ml. of 1.0 N aqueous hydrochloric acid is heated under reflux for a period of about three hours. The resulting two-phase system is cooledto a temperature of approximately C., and the layers are separated; The aqueous layer is extracted'with chloroform, and the chloroform extracts are combined with the original chloroform-dimethylformamidesolution. The combined organis layer is washed with an aqueous solution of sodium bicarbonate, and thechloroform and dimethyltormamide in the combined organic layer is replaced with ethyl ace- 14 tate by evaporation in vacuo. Petroleum ether is added and the resulting solution is subjected to a partition chromatogram using aqueous methanol as the stationary phase and benzene-chloroform as the moving phase to give 17m,2l-dihydroxy-16a-methyl-4-pregnei1e-3,1 1,20-trione.

A solution of 45 mg. of 3,ZO-bis-semicarbazido-17a,21- dihydroxy 16a-methyl-4-pregnene-3,11,20-trione 2l-acctate, 17 mg. of sodium borohydride, 1 ml. of tetrahydrofuran and 0.3 ml. of water is maintained at reflux temperature for approximately one hour. The reaction solution is cooled to about 15 C., and the excess sodium borohydride decomposed by the addition of a solution of 27 mg. of glacial acetic acid in 0.2 ml. of water. The tetrahydrofuran isevaporatcd in vacuo, and the residual material is extracted with ethyl acetate. The ethyl acetate extracts are Washed with a saturated salt solution, water, 5% aqueous sodium bicarbonate solution and again with water. The extracts are dried and the ethyl acetate evaporated in vacuo togive 3,20-bis-semicarbazido- 1 1,8, Hall-trihydroxy-16a-methyl-4-pregnene-3,20-dione.

A mixture of 60 mg. of 3,20-bis-semicarbazido- 11/8,17a,21 trihydroxy a methyl-4-pregnene 3,20- dione, 02 ml. of dimethylformamide, 0.6 ml. of chloroform, and 1.5 ml. of 1.0 N aqueous hydrochloric acid is heated under reflux for a period of about three hours. The resulting two-phase system is cooled to a temperature of approximately 15 C, and the layers are separated. The aqueous layer is extracted with chloroform, and the chloroform extracts are combined with the original chloroform-dimethylformamide solution. The combined organic layer is washed with an aqueous solution of sodium bicarbonate, and the chloroform and dimethylformamide in the combined organic layer is replaced with ethyl acetate by evaporation in vacuo. Petroleum ether is added and the resulting solution is subjected to a partition chromatogram using aqueous methanol as the stationary phase and benzene-chloroform as the moving phase to give 11 fi,17a,21 trihydroxy-l6u-methyl-4-pregnene-3,20- dione. The latter material is reacted with an excess of acetic anhydride in pyridine at room temperature for a period of about fifteen hours, and the crude acetylated product is recrystallized from ethyl acetate to give 11/3, 17a,21 trihydroxy-16a-methyl-4-pregnene-3,20-dione 2l-acetate.

To a cooled solution of 600 mg. of ll,B,l7 x,2l-trihydroxy 16a-methyl-4-pregnene-3,20-dione 21 acetate in 5.0 ml. of dry pyridine is added 0.15 ml. of phosphorous oxychloride, and the mixture is allowed to stand at room temperature for a period of approximately 15 hours. The reaction solution is evaporated in vacuo at a temperature of about 20 C. to a volume of 2-3 ml. Seventeen milliliters of water is added slowly to the concentrated solution, with'stirring, and the aqueous mixture is extracted with ethyl acetate. The combined ethyl acetate extracts are washed with water, then with dilute aqueous hydrochloric acid solution, again with Water, and then with a dilute aqueous sodium bicarbonate solution. The

washed ethyl acetate solution is dried, and the solvent is evaporated in vacuo; the residual material is triturated with ether, and the crystalline material is recrystallized from ethyl acetate-ether to give 17a,21-dihydroxy-16omethyl-4;9(11) pregnadiene-3,20-dione 21 acetate. A suspension of 330 mg. of :,21-dihydroxy-16a-methyl- 4,9(11)-pregnadiene-3,20-dione 21-acetate and 1.8 g. of N-bromosuccinimide in a mixture of 50 ml. of dioxane and 10 ml. of Water is cooled to 10 C. Then with stirring, 10' ml. of cold 1.0 N aqueous perchloric acid is added. The temperature of the reaction mixture is allowed to rise to 15 C. and is maintained at this point for about two and one-half hours during which time the solid material slowly dissolves. The resulting yellow solution is treated with 1.0 ml. of allyl alcohol to discharge the color and decompose the remaining N-bromosuccinimide, and the resulting solution is evaporated to a small volume in vacuo. The concentrated solution is diluted with water, and the aqueous mixture is extracted with ethyl acetate. The ethyl acetate extracts are Washed, dried and evaporated to dryness, and the residual material is crystallized from ethyl acetate-ether to give 9abromo l1,6,17u,21 trihydroxy l6a-methyl-4-pregnene- 3,20-done Zl-acetate.

A solution of 210 mg. of 9a-bromo-l1B,l7u,2l-trihy droxy-16tt-methyl-4-pregnene-3,ZO-dione Zl-acetatc and 240 mg. of potassium acetate in ml. of absolute ethanol is heated under reflux for two hours. The reaction mixture is cooled to room temperature, and evaporated in vacuo to a small volume. The concentrated aqueous mixture is extracted with three portions of ethyl acetate, and the combined ethyl acetate extracts are washed with Water, dried, evaporated to dryness in vacuo. The residual material is crystallized from ethyl acetate-ether to give 93,115 epoxy 17a,21-dihydroxy-16u-methyl-4-pregnene-3,20-dione 2l-acetate. To a solution of 83 mg. of anhydrous hydrogen fluoride in 4.7 ml. of ice-cold alcohol-free chloroform is added an ice-cold solution of 416 mg. of 98,1 1 B-epoxy-lhQl-dihydrox -16ct-methyl-4-pregnone-3,20-dione ZI-acetate. The solution is mixed thoroughly and maintained at 0 C. for two hours, at the end of which time 13.0 ml. of ice-cold aqueous sodium acetate is added, and the resulting mixture agitated vigorously. The layers are separated, and the chloroform layer is washed with water until free of acid, dried, and the chloroform evaporated in vacuo. The residual material is crystallized from acetone-petroleum ether to give 90c iluoro-llfi,17 x,2l-trihydroxy-l6a-rnethyl-4-pregnene 3,20-dione 2l-acetate. Fifty milligrams of 9a.- fluoro-l 1/5', l'7u,2 l-trihydroxy-16m-methyl-4-pregnene-3, 20- dione 21-acetate is dissolved in a mixture of 1.0 cc. of 1 N methanolic potassium hydroxide, and the solution is allowed to stand at room temperature for a period of about ten minutes. The solution is then acidified with acetic acid, the benzene is evaporated in vacuo, and the residual material is crystallized from ethyl acetate-ether to give 90 fiuoro-llfi,l7a,2l-trihydroxy 16a methyl-4- pregnene-3,20-dione.

A solution of 400 mg. of 9a-liuoro-l l,8,l7o,2l-trihydroxy-16ot-"nethyl-4-pregnene-3,ZO-dione ZI-acetate in 4 ml. of pyridine is added to the complex formed by the addition of 400 mg. of chromium trioxide to 4 ml. of pyridine. The mixture is swirled until thoroughly mixed and then allowed to stand at room temperature overnight. The reaction mixture is poured into water, and the aqueous mixture is extracted with ether and then twice with ethyl acetate. The combined ether and ethyl acetate extracts are Washed with dilute aqueous sulfuric acid at about 0 C., and then with water until neutral. The organic solvent layer is then dried, the solvents are evaporated therefrom in vacuo, and the residual crystalline material is purified by crystallization from ethyl acetate-ether to give Sa-iluoro-l7a,21-dihydroxy-16a-methyl- 4-pregnene-3,l1,20-trione Zl-acetate. Fifty milligrams of 9oc-fiuOrO-l7a,21 dihydroxy-l6a-methyl-4-pregnene-3,l1,- 20-trione 2l-acetate is dissolved in a mixture of 1.0 cc. of benzene and 1.0 cc. of 1 N methanolic potassium hydroxide, and the solution is allowed to stand at room temperature for a period of about ten minutes. The solution is then acidified with acetic acid, the benzene is evaporated in vacuo, and the residual material is crystallized from ethyl acetate-ether to give 9a-fluoro-l7oa2ldihydroxy-l 6a-methyl-4-pregnene-3 l 1 ,20-trione.

To a solution of 110 mg. of 17a,2l-dihydroxy-16 xmethyl-4-pregnene-3,11,20-trione 21-acetate in 6 ml. tbutanol, 0.01 ml. glacial acetic acid and 0.03 ml. of acetic anhydride is added 70 mg. of selenious acid (H SeO The mixture is heated to the boiling point overnight, another mg. of selenious acid is added, and the heating is continued for an additional 24 hours. The solution is decanted from metallic selenium and evaporated to an oil which is then dissolved in ethyl acetate. The ethyl acetate solution is washed with aqueous sodium bicarbonate and then with water until neutral, and dried. The solvent is evaporated from the dried solution to give an oil which is dissolved in benzene and adsorbed from this solvent on acid-washed alumina. The adsorbate is eluted with either-petroleum either and then with mixtures of ether and chloroform, increasingly rich in chloroform. The 4:6 ether-chloroform eluates are combined, evaporated to dryness, and the residual material recrystallized from ethyl acetate-ether to give 17a,21-dihydroxy-l6umethyl-l,4-pregnadiene-3,11,20-trione 21-acetate; M.P. 208-2l2 C. Fifty milligrams of l7a,2l-dihydroxy-16mmethyl-1,4-pregnadiene-3,11,20-trione ZI-acetate is dissolved in a mixture of 1.0 cc. of benzene and 1.0 cc. of l N methanolic potassium hydroxide, and the solution is allowed to stand at room temperature for a period of about ten minutes. The solution is then acidified with acetic acid, the benzene is evaporated in vacuo, and residual material is crystallized from ethyl acetate-ether to give 17,2l-dihydroxy-l6a-methyl-L4 pregnadiene 3,11,20- trione.

To a solution of mg. of llB,17a,2l-trihydroxy- 16 x-methyl-4-pregnene-3,20-dione 21-acetate in 6 ml. tbutanol, 0.01 ml. glacial acetic acid and 0.03 ml. of acetic anhydride is added 70 mg. of selenious acid. The mixture is heated to the boiling point overnight, another 50 mg. of selenious acid is added, and the heating is continued for an additional 24 hours. The solution is decanted from metallic selenium and evaporated to an oil which is then dissolved in ethyl acetate. The ethyl acetate solution is washed with aqueous sodium bicarbonate and then with Water until neutral, and dried. The solvent is evaporated fnom the dried solution to give an oil which is dissolved in benzene and adsorbed from this solvent on acid-washed alumina. The adsorbate is eluted with ether-petroluem ether and then With mixtures of ether and chloroform, increasingly rich in chloroform. The 4:6 ether-chloroform eluates are combined, evaporated to dryness, and the residual material recrystallized from ethyl acetate-ether to give l1B,l7oa,21-trihydroxyl6a-methyl-l,4-pregnadiene-3,20-dione 2l-acetate. Fifty milligrams of 11,8,l7a,21-trihydroxy-16a-methyl-L4-pregnadiene-3,20-dione ZI-acetate is dissolved in a mixture of 1.0 cc. of benzene and 1.0 cc. of 1 N methanolic potassium hydroxide, and the solution is allowed to stand at room temperature for a period of about ten minutes. The solution is then acidified with acetic acid, the benzene is evaporated in vacuo, and the residual material is crystallized from ethyl acetate-ether to give 1l;8,l7a,21-trihydroxy-lGot-methyl-1,4-pregnadiene-3,20-dione.

To a solution of mg. of 9oc-fillOIO17oz,21-dihydroxy-l6a-methyl-4-pregnene-3,l1,20-trione 21-acetate in 6 ml. t-butanol, 0.01 ml. glacial acetic and 0.03 ml. of acetic anhydride is added 70 mg. of selenious acid (H SeO The mixture is heated to the boiling point overnight, another 50 mg. of selenious acid is added, and the heating is continued for an additional 24 hours. The solution is decanted from metallic selenium and evaporated to an oil which is then dissolved in ethyl acetate. The ethyl acetate solution is washed with aqueous sodium bicarbonate and then with water until neutral, and dried. The solvent is evaporated from the dried solution to give an oil which is dissolved in benzene and adsorbed is eluted with ether-petroleum ether and then with mixtures of ether and chloroform, increasingly rich in chloroform. The 4:6 ether-chlorofiorm eluates are combined, evaporated to dryness, and the residual material recrystallized from ethyl acetate ether to give 9a-fluoro-l7a,2l-dihydroxy-16ot-methyl-1,4-pregnadiene 3,11,20-trione 2l-acetate. Fifty milligrams of 9tx-fiuoro-l7oa,21-dihydroxy-16amethyl-1,4-pregnadiene-3,11,20-trione 2l-acetate is dissolved in a mixture of 1.0 cc. of benzene and 1.0 cc. of 1 N methanolic potassium hydroxide, and the solution is allowed to stand at room temperature for a period of about ten minutes. The solution is then acidified with acetic cooled. It has 17 acid, the benzene is evaporated in vacuo, and the residual material is crystallized from ethyl acetate-ether to give 90:-

fluoro-17a,21-dihydroxy-16a methyl 1,4 pregnadiene- I 3,11,20-trione.

' To a solution of 100 mg. 9a-fiuoro-11p,17a,2l-

:trihydroxy l6a-methyl-4rpregnene-3,20-dione 21-acetate in v 6 m1. t-but-anol, 0.01 ml. glacial acetic acid and 0.03 ml.

of acetic anhydride is added 70 mgfiof selenious acid.

The mixtureis heated to; the boiling point 'overnight, an-

other 50 mg. of selenious acid is added, and'the'heating is continued for an .additiona1124 hours. The solu ition is decantedfrom metallic; selenium and evaporated from this solvent on acid-washed alumina. The adsorbateis eluted with ether-petroleum ether and then with mixtures of .ether 'and chloroform increasingly rich 1 in chloroform; The 4:6 ether-chloroform eluates are combined, evaporated to dryness, and the residual material recrystallized from ethyl acetate-ether to give 90ciluoro 11e,17u,21 trihydroxy 16oz methyl 1,4 pregnadiene-3,20-dione 21-acetate. Fifty milligrams of 904- fluoro 115,17a,21 tn'hydroxy 16a methyl 1,4 pregnadiene-3,20-dione 2l-acetate is dissolved in a mixture of 1.0 cc. of benzene and 1.0 cc. of 1 N methanolic potassium hydroxide, and the solution is allowed to stand at room temperature for a period: of about ten minutes. The solution is then acidified with acetic acid, the benzene is evaporated in vacuo, and the residual material is crystallized from ethyl .acet-ate ether to give 9uc-fi11010- 11 3,17 oc,21 trihydroxy l6cemethyl 1,4 pregnadiene- 3,20-dione.

In a. 500 ml. 3-neckfiask equipped with condenser, dropping funnel and nitrogen inlet are placed 20 g. of potassium hydroxide in 90 ml. of water, 100 ml. of methanol and 100 ml. of ether. A solution of 10 g. of N-methyl-N-nitroso-tosylamide in 50ml. of ether is placed in the dropping funnel. V

Diazomethane is generated by Warming the generation flask to 40-45 C. and cautiously adding the N-methyl-N- nitrosotosylamide-ether from the dropping funnel. Nitrogen is utilized to sweep the diazornethane into a solution of g. of 3a-acetoxy 16-pregnene-l1,20-dione in 100ml. of tetrahydrofuran and Y120 ml. of ether. The process is continued until the steroid solution remains yellow for several hours. The product, 3'a-acetoxy- 16oz,l7or-mcthyleneazorpregnane 11,20-dione largely precipitates from the reaction mixture. After 16 hours, the

mixture is filtered, washed with ether and dried in air.

. Yield about 14 grams, M.P. 186-190" C.

A manometer and 12-liter surge flask are in the line between the reaction flask and pump trap. When the bath temperature reaches 180 C. the 30t-3C6tOXY-l60t,170t,

methyleneazo-pregnane-l1,20-dione begins to melt with evolution of nitrogen. Themaximum pressure reached is 83 mm. After 10 minutes at 180-182 C. the melt is OH on a 249 E percent 191 and is taken up in about 150 ml. of acetone,

filtered through diatomaceous earth, concentrated to about 100 ml., and ether is slowly added to the boiling solution until crystallization occurs. These crystals of 30aacetoxy-16-methyl-16-pregnene-11,20-dione Weigh about 19.0 g., M.P. 165-168" C. p

' A solution of 20.0 g. of 3a-acetoxy-16methyl-16-pregnene-11',20-dionedissolved in 600 ml. of methanol, is cooled to 18 C., and 80ml. of 30% hydrogen peroxide followed by 80 ml. of 2.5 N sodium hydroxide are added.

Considerable material precipitates from solution, but all redissolves on stirring the reaction mixture at -30 C. for 40 minutes. .The solution is kept at 1520 C. for 18 hours at which time the ultraviolet maximum at 249 has completely disappeared. Then 600 ml. of saturated salt water is slowly added, the crystalline precipitate is filtered, washed with Water, and dried in air: and in vacuum. The 16u,17a-epoxy 3oz hydroxy-16fl-methylpregnane-11,20-dione thus formedweighs about 17 g.; M.P. 176-1'7 7 C.; hexagonal prisms, M.P. 178-180. C., :from acetone-ether;

To a solution of 2.69 g. of 16a,17a-epoxy-3a-hydroxy- 16B-methyl-pregnane-11,20-dione in 55 dioxane is added 27 ml. of 2 M aqueous perchloric acid. The clear solution is kept at 25-30 C. for 65 hours.

after 30 minutes. The precipitate, containing a mixture of 3 a,17 a dihydroxy-l6 -methy1-lS-Pregnene-I1,20-dione and 3 or,17a-dihydr0xy-16-methylene-pregnane-l 1,20-dione is Washed with Water, and dried in air and finally at 50 C. in vacuum. Yield: approximately 2 g.; M.P. sintering at about 150 C., melting at 158-167 C. The relative proportion of 3oz, 17a-dihydroxy-16-methyl-l5-pregnene- 11,20 dione and 30,l7u dihydro-xy-lG-methylene-preg- "taken to dryness and crystallized from ether; a mixture of tained; weight about 3 3 a, 17 a-dihydroxy- 1 6a-methyl-pregnane-1 1,20 di one and 3oz,17a-dihydroxy-16/3-methyl-pregnane-11,20-dione is obv g., sintering at 150 C. melting at 166482 c. 7 p

' max.

The product consists of 3a,17a-dihydroxy-16a-methylpregnane-11,20-dione and 3 x,17a-dihydroxy-16p-methylpregnane-11,20-dione in the. ratio ca. 7:3 as determined by the amounts of end product isolated below.

One gram of this hydrogenation product containing 3a,17a-dihydroxy-16a-methyl-pregnane-11,20 dione and 3a,17a,dihydroxy-16;8-methyl pregnane 11,20 dione, is chrom-atographed on 100 g.-of activitated magnesium silicate. The 100% chloroform eluates give 3a,17a-dihydroxy-16e-methyl-pregnane-11,20-dione, M.P.. l88-19l C. The 5% methanol-chloroform eluates give 3a,17a-

dihydroxy-l 6B-methyl-pregnane 11,20 dione, hexagonal plates from benzene-ethybacetate;M,P. 192-197 C.

A- solution of 3.50 g. (9.7 rnillimols) of 3a,17a-dihydroxy-16fl-methyl-pregnane-11,20-dione in 40 ml. of

chloroform is warmed to 4045 C. A solution of 1.76 g. (11 millimol) of bromine in 25 ml. of chloroform is added dropwise to the stirred solution such that the color is not darker than pale yellow (ca. 2-drops/sec., total time1 hour). The nearly colorless solution is cooled to 20 C. and 200 ml. of ether is added; The mixture is extracted with excess cold 5% potassium bicarbonate solution, sodium bisulfite solution, and water, and dried over magnesium sulfate. The colorless residue after re moval of solvent, 21- bromo-3a,17a-dihydroxy-16/5methylgrams) gives a positive Cold water ml.) is added, the slurry chilled to 8 C. and filtered The stirrecl'mixture 19 extracted into ethyl acetate, and the organic extract dried over magnesium sulfate to give about 4 grams of a colorles foam that partly crystallizes from acetone-ether to give 30 ,170t,21 trihydroxy 16p methyl-pregnane-11,20- dione 21-acetate. I

To a solution of 30,17a,21-trihydroxy-16fl-methyl-pregnane-l1,20-dione 2l-acetate (4.0 g.) in 100 ml. t-butanol and 20 ml. of water cooled to l15 C., is added 3.5 g. N-bromo-succinimide; The suspension is stirred at C. until all the N-bromosuccinimide has dissolved (90 minutes). The reaction mixture is kept at 2 C. for about sixteen hours and at 25 C. for 2 hours. Sodium sulfitesolution is added to destroy bromine and the mixture concentrated on the Water pump to a low volume. A granular precipitate forms; water is added, the precipitate filtered and washed with water; chromatography on neutral alumina and elution with mixtures of chloroform and benzene gives 17a,21-dihydroxy-16,8-methyl-pregnane-3,11,20-trione 21-acetate, MP. 210213 C.

To a stirred solution of 585 mg. of 17a,2l-dihydroxy- 6fl-methyl-pregnane-3,11,20-trione 21-acet-ate in 10 ml. of acetic acid and 8 ml. of chloroform kept at 10 C. is added slowly 230 mg. of bromine in 6 ml. of chloroform. After addition is complete, 1.2 g. of sodium acetate in 7 ml. of cold Water is added and the mixture is extracted with chloroform. The chloroform extract is Washed with dilute potassium bicarbonate, water and dried over sodium sulfate. The residue is t riturated with ether to give 480 mg. of crystalline 4-bromo-17u,2l-dihydroxy- 16fl-methyl-pregnane-3,l1,20-trione 2l-acetate, M.P. 165- 170 C., dec. J

To 583 mg. of of 4-bromo-l7a,21-dihydroxy-16(3-methyl-pregnane-3,11,20-trione 21-acetate in ml. of acetonitrile under nitrogen is added a slurry of 600 mg. of semicarbozide hydrochloride and 410 mg. sodium bicarbonate in 4 ml. of Water. After 2 hours, the acetonitrile is removed in vacuo, water is added and about 540 mg. of crystalline 3-semicar-bazone of 17a,21-dihydroxy-l6,8- methyl-4-pregnene-3,11,20-trione 21-acetate filtered, wash ed with water and dried.

540 mg. of the 3-semicarbazone of l7oc,2l-dihydroxy- 16/8-methyl-4-pregnene-3,11,20-trione 21-acetate is dissolved in 20 ml. of acetic acid, 1.5 ml. of pyruvic acid and 5 ml. of Water. After 18 hours at C., water is added and the mixture extracted with chloroform. The chloroform extract is washed with aqueous potassium bicarbonate, water and dried over sodium sulfate. Removal of solvent gives crude 17a,2l-dihydroxy-16fi-methyl-4- pregnene-3,11,20-trione 21-acetate which is purified by chromatography on neutral alumina and crystallization from acetone-ether (hexagonal plates). The pure material has Ml. 226-232 C. 100 mg. of 17ot,21-dihydroxy- 16/3-methyl-4-pregnene-3,l1,20-trione 21-acetate in 3 ml. of methanol is treated with 0.1 g. of potassium bicarbonate in 1 ml. of Water, the methanol is evaporated in vacuo, and the residual material is extracted with ethyl acetate. Evaporation of the ethyl acetate solution gives crystalline 17u,21-dihydroxy 16/8 methyl-4-pregnene- 3 ,l1,20-trione.

To a stirred solution of 500 mg. of 17a,21-dihydroxyl 6 3-methyl-4-pregnene-3,l1,20-trione 2l-acetate in 12.5

ml. of methanol and 3 ml. of dimethylformamide kept under nitrogen is added a slurry of 680 mg. of semicarbazide hydrochloride and 370 mg. of sodium bicarbonate in 1 ml. of water. The stirred mixture is refluxed 3 /2 hours and maintained at 45 C. for 17 hours. It is then cooled to 20 C. and 50 ml. of 50% saturated aqueous sodium chloride is added. After 2 hours at 0 C. the precipitate of 3,20-bis-semicarbazido-l7a,2l-dihydroxy- 16,8-methyl-4-pregnene-3J1,20-trione 21-acetate is filtered, washed with water until free of chloride ion and dried in air.

To a stirred solution of 600 mg. of 3,20-bis-semicarbazido 1 711,21 dihydroxy 16,8 methyl 4 pregnene- 3,11,20-trione 21-acetate in ml. of tetrahydrofuran and 11 ml. of water under nitrogen is added 200 mg. powdered sodium borohydride. The stirred suspension is refluxed 45 minutes and then cooled to 15 C. Aqueous acetic acid (3 ml. of 30%) is added cautiously and most of the tetrahydrofuran is removed in vacuum. Addition of 5 ml. of methanol and 5 ml. of water induces the prod not to crystallize. Following addition of 10 ml. of a saturated sodium chloride solution and aging at 0 C. the product 3,20-bis-semicarbazido-11,B,17a,21-trihydroxy- 16B-methyl-4-pregnene-3,20-dione is filtered, Washed with Water, and dried in air.

To a solution of 510 mg. of reduced 3,20-bis-semicarbazido-l1B,17a,2l-trihydroxy-16fl methyl-4-pregnene- 3,20-dione in 5 ml. of acetic acid is added 1.20 ml. of water and 0.50 ml. of pyruvic acid. The solution is kept at 25 C. for eighteen hours. Water (20 ml.) is added, and the mixture is extracted thoroughly with chloroform. The chloroform extract is dried over magnesium sulfate and taken to dryness. The residue is crystallized from acetone-ether to give pure 116,17u,21-trihydroxy-l6fimethyl-4-pregnene-3,20-dione. A solution of mg. of 1l,8,17u,21 trihydroxy 16fi-methyl-4-pregnene-3,20- dione in 1.0 ml. of pyridine and 0.5 ml. of acetic anhydride is prepared. After 18 hours at 25 C., the solution is taken to dryness in vacuo and the solid residue purified by crystallization from acetone-ether to give 11,B,l7oc,2ltrihydroxy-16,8-methyl-4-pregnene-3,20-dione 2l-acetate.

A solution of 400 mg. of 1lfi,17a,21-trihydroxy-16l3- methyl-4-pregnene-3,20-dione 21-acetate in 2.0 ml. dimethyl formamide, 0.8 ml. of pyridine and 0.4 ml. of methanesulfonyl chloride is kept at 75 'C. for one hour. The mixture is cooled, water added, and the precipitate filtered, washed with Water and dried in air. The desired l7a,21-dihydroxy-16fl-methyl-4,9 1 1) pregnadiene-3,20- dione 21-acetate is purified by chromatography on alumina (20 g.) and elution of the column with benzene. Crystallization of material eluted by benzene gives pure 17a,21-dihydroxy 16/3 methyl 4,9(11) pregnadiene- 3,20-dione 21-acetate.

. To a mixture of 620 mg. of 17u,21-dihydroxy-16/3- methyl-4,9( 11)-pregnadiene-3,20-dione 21-acetate and 330 mg. of N-bromosuccinimide in 10 ml. of dioxane and 3.2 ml. of water cooled to 10 C. is added 1.8 ml. of cold 1M aqueous perchloric acid. The mixture is stirred at 15 C. for 3 hours. Excess N-bromosuccinimide is destroyed by addition of aqueous sodium thiosulfate and most of the dioxane is removed in vacuo. About 30 ml. of water is added and crystalline bromohydrin, 9a-bromo-11,8,17a,21-trihydroxy-16,8 methyl-4- pregnene-3,20-dione 21-acetate, is filtered, washed with water, and dried in air.

To a stirred solution of 100 mg. of the 9oa-br0mo-1L8, l7or,2l-trihydroxy-l6fl-methyl-4-pregnene 3,20 dione 2l-acetate in 3 ml. of tetrahydrofuran and 1 ml. of methanol under nitrogen is added 1.02 ml. of 0.215 N methanolic sodium methoxide. After 10 minutes at 25 C., 0.2 ml. of acetic acid is added and the methanol removed in vacuo. The residue is acetylated with 1.00 ml. of pyridine and 0.5 ml. of acetic anhydride at 60 C. for 70 minutes. The mixture is taken to dryness in vacuo, water added, and the product extracted into chloroform. The residue is crystallized from ether-acetone to give pure 9,8,11l3 epoxy-17m,2l-dihydroxy-16fi-n1ethyl4-preg nene-3,20-dione 21-acetate.

To a solution of 200 mg. of 9fi,1lB-epoxy-l7a,21-dihydroxy-l6fl-methyl-4-pregnene-3,20-dione 21-acetate in 2 ml. of chloroform and 2 ml. of tetrahydrofuran in a polyethylene bottle at 60 C. is added 2 ml. of a 2:1 (by weight) mixture of anhydrous hydrogen fluoride and tetrahydrofuran. After 4 hours at -10 C. the mixture is cooled to -60 C. and cautiously added to a stirred mixture of 30 ml. of 25% aqueous potasisum carbonate and 25 ml. of chloroform kept at 5 C. The aqueous phase is further extracted with chloroform and the latter phase washed with Water and dried over magnesium sul- After 15 minutes, 0.2 ml. of acetic acid in 1 ml. of water is addedand the mixtureconcentrated nearly to dryness. The residue is taken up in Lefthyl acetate' andthe ethyl acetate solution is washed witlr'water, dried over magnesium sulfate, and concentrated to dryness. Crystallization ofthe residue from ethyl acetate gives pure 9asiuin bicarbonate in aqueous methanol to give S a-chloro- 11/3,l7oz,2l-trihydroxy-l6B-methyl-4-pregnene-3,20 dione.

To. a stirred solution of 682 mg. of 17a,21-dihydroxy- 7 22 hydrochloric acid and water and dried over magnesium sulfiate. It is then treated with activated charcoal and concentrated to dryness. Crystallization of the residue from acetone-ether gives pure 11fi,17ot,21-trihydroxy-16B- methyl-1,4-pregnadienc-3,20-dione ZI-acetate. 100 mg. of 1l[3,l7oc,21 trihydroxy --1GB-methyl-1,4-pregnadiene- 3,20-dione 21 acetate in.3 ml. ofmethanol is treated with i 0.1 g. of potassium bicarbonate in ml. of water under 16/5' methyl-pregnane-3J1,2O-trione 2l-acetate in 20 ml.

of chloroform and 2.25 ml. of acetic acid maintained at 20 C. is added dropwise one-half of a solution of 540 mg. of bromine in 2 ml. of chloroform and 3 ml.- of acetic acid. The mixture is warmed to 0 C. and the remainder of the bromine added. Sodium acetate (0.4 g.) in 2 ml. of water is added followed by 20 mg. of sodium sulfite. The mixture is concentrated in vacuo to remove the chloroform and 20 ml. of water is added. The white powdery precipitate of 2,4-dibromo-17u,21-

dihydroxy-l6p-methyl-pregnane-3,1 1,20-trione 2 l-acetate is filtered, washed with water and dried in air. Yield: 920 mg. M.P. 122.130 C., dec.

To a solution under nitrogen of 900 mg. of the 2,4- dibromo 170:,21 dihydroxy-16fl-methyl-pregnane-3,l1, ZO-trione 21-acetate in 5 ml. dimethyl formamide is added 200 mg. of sodium bromide. After lhour at 25 C., 1 ml. of, dimethylaniline is added and the mixture maintained at 135 C. for 2 /2 hours. The mixture is cooled, added dropwise to dilute hydrochloric acid, and solid crude. P oduct filtered, washed with dilute hydrochloric acid, water and dried in air. Treatment withcharcoal, followed by crystallization from acetone gives 17oz,21.-di-

hydroxy 16B methyl-1,4-pregnadiene-3,11,20-trione 21- 1 1.0 g. of l7tx,2l-dihydroxy-16/3-methyl-1,4-pregnadiene- 3,11,20-trione 2l-acetate in 30 ml. of methanol is treated With 1 g. of potassiumbicarbonate in 10 ml. of water under nitrogen at reflux temperature for 7 minutes. mixture is cooled, neutralized with 1 ml. of acetic acid in 10 ml. of water, the methanol removed in vacuo and the product extracted into ethyl acetate. Removal of the ethyl acetate gives crystals (about 0.9 g.) of the desired 17a,21-dihydroxy 16B methyl-l,4-pregnadiene-3,11,20- trione; M.P. 195200 C. a

To 100 mg. of 11,B,1'Zu,21-trihydroxy-16,8-methyl-4- pregnene-3,20-dione ZI- aoetate in 5 ml. of acetic acid is added 50 mg. of selenium dioxide. The mixture is reliuxed under nitrogen 18 hours, 50 mg. of selenium dioxide is added and the mixture refluxed an additional 24 hours. The mixture is filtered, and the filtrate taken to dryness. The residue is taken up in ethyl acetate and washed successively with aqueous sodium bicarbonate,

ammonium sulfide,'dilute ammonia water, water, dilute The- ' bonate in aqueous methanol to '=dihydroxy-16,8-methyl4-pregnene-3, 1 1,2Q-trione.

j i-pregn-ene-3,2O-dione ZI-acetate is reacted with chrornium rtrioxide-pyridine complex to-form 9ot-ch1-oro-l7a, ZI-dihydroXy-I6B-met-hyl-4-pregnene-3,l1,20 trione 21- nitrogen at reflux tempenature for 7 minutes. The mixture is cooled',;neutralizedwith 0.1 of acetic acid in 1 ml. of water,-;the methanol is evaporated in vacuo and the residual material is extracted with ethyl acetate.

21-trihyd1'oxy-16,6-methyl-l,4 pregnadiene-3,ZO-dione.

In a'similar manner, wil ing. of 9a-fluoro-llfi,l7ot,2ltrihydroxy-l65-methyl-4-pregnene-3,ZO-dione 21 acetate is treated with selenium dioxide t-o produce the correspending 9a fluono-l'l/3,=17a,21-trihydnoxy-16,8-methyl-1,4- pregnadiene- 3,20-dione ill-acetate. T his compound is hydrolyzed with potassium bicarbonate in aqueous methanol in accordance with the pnocedure described in the preceding panag raph to give 9ot-fluoro-l1 8,l7ct,2latrihydroxy-16fl-metzhyl-1,4-pregn-adiene-3,2O dione. Similarly, mg. of 9u-ch1oro-1l,8,=l7ot,21-trihydroxy-16B- methyl-4-pregnene-3,2fi dione 21-acetate is treated with selenium dioxide to produce 9vx-chloro-11fi,17ot,2ltrihydnoxyB-methyll,4-pregnadiene-3,20-dione 21 acetate, which is hydrolyzed by treatment with potassium bicarbonate in aqueous methanol to give 9a-'chloro-1 1fi,17u,2ltrihydr-oxy--methyl-1,4-pregnadiene-3,ZO-dione.

A solution of 100 mg. of 9ot-fluoro-11B,17u,21-trihydroxy-16/3-methyl-1,4-pregnadiene-3,ZO-dione 21' acetate in 1ml. of pyridine is added to the complex formed by the addition of 100 mg. of chromium trioxide to 1 ml. of pyridine. The mixture is swirled until thoroughly mixed and then allowed to stand at room temperature overnight. The reaction mixture is poured into water, and the aqueous mixture is extracted with ether and then twice with ethyl acetate. The combined ether and ethyl acetate extracts are washed with dilute aqueous sulphuric acid at about.0 C., and then with water until neutral. The organic solvent layer is then dried, the solvents are evaporated therefrom in vacuo, and the residual crystalline material ispuirified by crystallization from ethyl ace- (rate-ether to give 9o-fiuono-17a,2l-d-ihydroxy-16,8-methyl- =1,4-pregnadiene-3,l1-20-trione 21-.aoetate. This compound is hydrolyzed by treatment with potassium bicarbonate in aqueous methanol to form 9a-fillO1O-17a,2l-dihydroxy-lGui-methyl-1,4-pregnadiene-3,1 1,2O-trione.

Ina similar manner, 9a-chloro-l lfl,17a,21-trihydroxy- 16,8-methy1-1,4-pregnadiene-3,ZO-dione 21 acetate is reacted with chromium trioxide-pyridine complex to give 9o -chloro-17ot,21-dihyclroxy-16fi-methyl 1,4 pregnadiene 3,11,20 trione 21 acetate, which is hydrolyzed by treatment with potassium bicarbonate in aqueous methhydroxy-l6fl-methyl-4-pregnene-3,1:1,20-trione ZI-tacetate WilLCl'l is hydrolyzed by treatment with potassium bicarproduce 9ot-fluoro-17o ,'21-

Similarly, 9u-chloro l lfi,17a,21-trihydroxy-16 8-methylacetate which is hydrolyzed bytreatment with potassium bicarbonate in aqueous methanol to produce 9ot-chloro- 17u,2l-dihydroxy-16 3-methyl-4-pregnene-3,1 120-trione.

1 0 grams of 17a,21-dihydroxy-16ot-methyl-4-pregnene- 3,11,20-trione which can bepreparred as described hereinabove, are dissolved in 400 cc. of chloroform. Tothis solution is added a'mixture of 100 cc. of concentrated aqueous hydrochloric acid and 100 cc. of 37% aqueous formaldehyde solution. The resulting mixture is stirred for a period of approximately 3 days at room temperature. The chloroform layer is separated, washed with sodium bicarbonate solution until neutral, then with water, dried over anhydrous sodium sulfate, and evaporated to dryness in vacuo. The residual crude material is purified by crystallization from a mixture of methylene chloride and methanol to give 17a,20,20, 21-bismethylenedioxy-l6amethyl-4-pregnene-3,ll-dione.

lOgrams of 170:,20,20,21-bismethylenedioxy-16u-methyl-4-pregnene-3,1l-dione are dissolved insSOO cc. of benzene, to the solution is added 25 cc. ethylene glycol and 1 gram of p-toluenesulfonic acid, and the resulting mixture is heated under reflux fora period of approximately 15 hours. The reaction solution is cooled, washed with aqueous sodium bicarbonate solution, dried over anhydrous sodium sulfate, and evaporated to dryness in vacuo. The residual crude material is absorbed from its solution in benzene on 250 g. of acid-washed alumina, and the resulting adsorbate is eluted utilizing mixtures of ether-petroleum ether to give 3-ethylenedioxy-17a,20,20, 2 l-bisrnethylenedioxy-16a-methy1-5-pregnene-1 l-one.

A mixture of 10 grams of 3-ethylenedioxy-l7ot,20,20, 2l-bismethylenedioxy-l6oz-methyl-5-pregnene-1l-one, 4.3 g. perbenzoic acid and 230 cc. benzene is allowed to stand at room temperature for a period of about 2 days. A portion of the epoxide product, which precipitates, is separated from the reaction solution by decantation, and the decanted solution is washed with saturated aqueous sodium bicarbonate solution and then dried over anhydrous sodium sulfate. The dried solution is 'then evap-' orated in vacuo, and the residual epoxide product is combined with the epoxide product originally separated from the reaction mixture.

The mixed epoxides are dissolved in 200 cc. of substantially anhydrous formic acid and the resulting solution is allowed to stand at room temperature for a period of about 2 /2 hours. The reaction solution is poured into water, and the aqueous mixture is extracted with chloroform. The chloroform solution is washed with saturated aqueous sodium bicarbonate solution, dried over anhydrous sodium sulfate, and evaporated in vacuo to give a mixture of 17a,20,20,-2l-bismethylenedioxy-S-hydroxy- 16a-methyl-6-formyloxy-pregnane-3,1l,dione and l7cz,20, 20,21 bismethylenedioxy 6 hydroxy 16;: methyl-5- formyloxy pregnane-B, ll-dione.

' This mixture of 5-formyloxy and -formyloxy derivatives is dissolved in about 850 cc. of methanol, to this so lution is added a solution containing about 17 grams of potassium hydroxide in 80 cc. of Water, and the resulting mixture is heated under reflux in a nitrogen atmosphere for a period of approximately /2 hour. The reaction solution is cooled, neutralized with about 23 cc. of acetic acid and evaporated in vacuo to a small volume. The concentrated solution is poured into water and the aqueous mixture is extracted with chloroform. The chloroform extract is washed with aqueous sodium bicarbonate solution, dried over anhydrous sodium sulfate, and evaporated to dryness in vacuo. The residual material is crystallized from methanol to give 17ct,20,20,2l-bismethylenedioxy- 16e-methyl-allopregnane-3,6, l l-trione.

A solution containing about 5 grams of 17OL,20,20,21- bismethylenedioxy 16o: methyl allopregnenefi,lil-trione and :1 g. of p-toluenesul fonic acid in 120 cc. butanone dioxolane is heated under reflux for a period of about 10 minutes. The reaction solution is cooled to about 0- C., diluted with chloroform, and the resulting solution is washed with aqueous odium bicarbonate solution and dried over anhydrous sodium sulfate. The resulting dried solution is evaporated to dryness in vacuo. and the residual material is slurried with ether, recovered by filtration and then dried to give 3-ethylenedioxy-l7oc,20,20,21- bismethylenedioxy 160a methyl-allopregnane-6,1l-dione.

' A solution containing about 5 grams of 3-eth'ylenedioxy- 17oc,20,20,21 bisrnethylenedioxy 16oz methyl allopregnane-6,1l-dione in 70 cc. of benzene is added, with stirring over a period of approximately 10 minutes, to an ethereal solution containing the methyl magnesium iodide, prepared from 5 cc. of methyl iodide and 0.5 grams of magnesium turnings, dissolved in 50 cc. of ether. The reaction mixture is allowed to stir for an'additional /2 hour period, and the resulting solution is decomposed with 70 cc. of water. About 1710 cc. of benzene is added to the aqueous mixture and the layers are separated. The aqueous layer is extracted with two 109 cc.-portions of chloroform, and the organic layers are combined, dried over anhydrous sodium sulfate, and evaporated to dryness in vacuo to give 3-ethylenedioxy-17a,20,2(),2l-bis- :methylenedioxy-flhydroxy 604,16 dimet-hyl-allopregmane-=1 l-one.

A solutioncontaining 5.5 cc. of freshly distilled thionyl chloride in 26 cc. of cold anhydrous pyridine is added dropwise, with stirring, to a solution of 5.0 g. of 3-ethylenedioxy-l7a,20,20,21 bis methylenedioxy-6fi-hydroxy- 6a,16u-dimethyl-allopregnene-1l-one in 32 cc. of anhydrous pyridine,.while maintaining the temperature of the reaction mixture at approximately 40 C. The reaction solution is stirred for an additional 30 minute period following the addition of the thionyl chloride reagent, and the reaction mixture is then cooled to about 0-5 C. and poured into 180 cc. ice water. The aqueous mixture is extracted with chloroform, and the chloroform extract is neutralized, dried over anhydrous sodium sulfate, and evaporated to dryness in vacuo. The residual material, dissolved in benzene, is chromatographed on grams of acid-washed alumina; elution of the resulting adsorbate with mixtures of petroleum ether and ether gives 3-ethylenedioxy-17a,20,20,21 bismethylenedioxy 6,1604 dimethyl-5 -pregnene-1 l-one.

A solution containing about 5 grams of 3-ethylenedioxy-l7a,20,20,2l-bismethylenedioxy 6,16a dimethyl- S-pregnene-ll-one, 500 cc. of anhydrous acetone and about 0.5 g. of p-toluenesulfonic acid monohydrate is allowed to stand for a period of about 15 hours. The reaction solution is poured into water, and the aqueous mixture is extracted with chloroform. The chloroform solution is washed with aqueous sodium bicarbonate solution, dried over anhydrous sodium sulfate, and the chloroform solution is evaporated to dryness in vacuo. The residual material is crystallized from benzene to give 17a,20,20,2l -bismethylenedioxy 611,160: dimethy1-4 pregnene-3,1l-dione.

About 0.1 gram of l7a,20,20,2l-bismethylenedioxy- 60c,l6oc-dilnethYl-4-Pfflgl16n6-3,ll-diOIlB is suspended in 18 cc. of 50% aqueous acetic acid, and the suspension is carefully purged with nitrogen, following which the suspension is heated on a steam bath under nitrogen for a period of about 8 hours and then allowed to stand at room temperature for an additional period of 15 hours. The reaction solution is evaporated to dryness in vacuo to give 17 0c,2I-dihYCiIOXY-Ga,l6oc-dimellllyl 4 pregnene- 3,11,20-trione. The material is dissolved in 20 cc. of chloroform, and the chloroform solution evaporated to dryness in vacuo. To the residual dry material is added 1 cc. of pyridine and 1 cc. of acetic anhydride, and the resulting mixture is heated on a steam bath for a period of about 15 minutes. The reaction solution is cooled to about O5 C. poured into ice water. The aqueous suspension is extracted with chloroform, and the chloroform extract is washed with aqueous sodium bicarbonate solution, dried over anhydrous sodium sulfate, and evaporated to dryness in vacuo. The residual material, dissolved in 30 cc. benzene, is adsorbed on 5 grams of acidwashed alumina; the adsorbate is washed with 50 cc. ether. The adsorbate is then eluted with 250 cc. of a 5:5 mixture of ether-chloroform, and then with 50 cc. of a 3:7 mixture of ether-chloroform. The eluates are in vacuo.

25 evaporated in vacuo, and the residual material is crystallized from methanol to give l7a,2l-dihydroxy-6a,16w. dimethyl- 4-pregnene-3,l1,20-trione 2l-acetat'e.

- In accordance with the foregoing procedures, but start-' ing with 17 :,21 dihydroxy 16B methyl 4.,- pregnene- 3,11,20-trione, there are obtained the corresponding 17a,21-dihydroxy-6a,16,8;dimethyl 4 pregnerie-3,11,20- trione and its Zl-acetate. t

To a solution of 100 mg. of 176:,2l-dihydroxy-6a,l6 xdimethyl-4-pregnene-3f1IQZO-trione ZI-acetate, in 6 ml. .t-butanol, and 0.01 ml. of glacial acetic acid is added 70 mg. of selenium dioxide. reflux in a nitrogen atmosphere for a-periodiof about '6' The mixture is heated under hours.

' zene solution is evaporated in vacuo. The residual material thus obtainedv is separated using partition chromatography on diatomaceous silica (Supercel) to give 1706,21-

dihydroxy-6a,l6 x dimethyl 1,4 pregnadiene-B,1l,20- trione 2l-acetate. 40 mg. of 17a,2ldihydI'OXy6a,16oc dimethyl-1,4-pregnadiene-3,11,20-trione 2l-acetate is dissolved in a mixture of 1.0 cc. of benzene and 1.0 cc. of 1 N methanolic potassium hydroxide, and the solution is allowed to stand at room' temperature for a'period of "about minutes. The solution is then acidified with acetic acid; then benzene is evaporated in vacuo, and the residual material is purified by crystallization to give 17a,2l-dihydroxy-6u,l6m dimethyl 1,4 pregnadiene- 3,11,20-trione.

Similarly, in accordance with the foregoing procedure,

but starting with 17a,21-dihydroxy-6a,16B-dimethyl-4- pregnene-3,11,20-trione ZI-acetate, there are obtained the corresponding 17,21-dihydroxy 60;,16/3 dirnethyl 1,4- pregnadiene-3,l1,20-trione, and its 2l-acetate.

About 5 grams of 3-ethylenedioxy-=17u,2,20,2l-bismethylenedioxy-6,l6oa-dimethyl-5-pregnene 1l-one, which can be prepared as described'hereinabove, is dissolved in about 85 cc. of benzene. This solution is added to a stirred suspension containing 5 grams of lithium aluminum hydride in one liter of anhydrous ether, and the re-v s'ulting suspension is heated under reflux for a periodjof about 4 hours. The reaction mixture is allowed to cool to about room'tempe-rature, and approximately 33 cc. of ethyl acetate is added to destroy excess lithium aluminum hydride. The rcsulting mixture is admixed with about 100 cc. of water, the ethereal layer is decanted, and the aqueous layer diluted with an additional 200 cc. of water, is extracted with chloroform. The ether and chloroform layers are combined, and the combined organic layers are dried over anhydrous sodium sulfate and evaporated to dryness in vacuo.

of acid-washed alumina, and the resulting adsorbate is The residual material, dissolved in about 250 cc. of benzene, is adsorbed on about 150 grams 25 About 011 gram of l7a,20,20,2 1-bismethylenedioxyllB-hydroxy-6a,ldot-dimethyl 4 pregnene-3-one is suspended in 18 cc. of 50% aqueous acetic acid, and thesuseluted utilizing mixtures of petroleum ether-ether. This eluate is evaporated in vacuo, and the residual material is purified by crystallization from ether to give 3-ethylenedioxy-l7rx,20,20,2l bismethylencdioxy-1lfi-hydroxy-6,160idimethyl-S-pregnene.

A solution containing about 5 grams of 3-ethylenedioxy 17a,20,20,21 bismethylenedioxy 11,8 hydroxy- 6,l6m-dimethyl-5-pregnene, 500 cc. of anhydrous acetone and about 0.5 g. of p-toluenesulfonic acid monohydrate .is allowed to stand for a period of about 15 hours. The

The

. pointa precipitate appears; the resulting mixture is then droxy-da,16a-dimethyl-4-pregnene-3,20-dione pension is carefully purged with nitrogen, following which the suspension is heated on a steam bath under nitrogen for a period of aboutS hours and then allowed to stand dryness in vacuo. To the-residualdry material is added. 1 cc. of pyridine and 1 cc. of acetic anhydride, and the resulting mixture is heated on a steam bath for a period of about 15 minutes. The reaction solution is cooled to about 0-5" C. and poured into ice water. The aqueous suspensionis extracted with chlorofornn and the chloroform extract is washed with aqueous sodium bicarbonate solution, dried over anhydrous sodium sulfate, and evaporated in vacuo. The residual material, dissolved in 30 cc. benzene, is adsorbed on 5 grams of acid-washed alumina; the adsorbate is washed with 50 cc. ether. The adsorbate is then eluted with 250 cc. of a 5:5 mixture of etherchloroform, and then with 50 cc. of a 3:7 mixture of ether-chloroform. The eluates are evaporated in vacuo, and the residual material is crystallized from methanol to give llfl,l7a,2l-trihydroxy 6oi,1-6oe dimethyl-4-pregnene-3,20-dione ZI-acetate. I

In accordance with the foregoing procedure, but starting with 3-ethylenedioxy-17 x,20,20,2l-bismethylenedioxy- 6,16/3-dimethyl-5-pregnene-l l-one (which is obtained as an intermediate in the preparation of l7a,2l-dihydroxy- 60,16fl dimethyl 4 pregnene-3,l1,20-trione 2l-acetate herein described above), there are obtained the corresponding 1 1 ,8,l7a,21 trihydroxy-Ga,l6fi-dimethyl-4-pregnene-3,20-dione and its 2l-acetate.

To a solution of mg. of 11,B,17a,21-'trihydroxy- 6a,16or-dimethyl-4-pregnene-3,20-dione Zl-acetate, in 6 ml. t-butanol, and 0.01 ml. of glacial acetic acid, is added 70 mg. of. selenium dioxide. The mixture is heated under reflux in a nitrogen atmosphere for a period of about 6 hours. The reaction solution is filtered and evaporated in vacuo. The residual material is dissolved in benzene, and the benzene solution is Washed with an aqueous sodium bicarbonate solution and dried over anhydrous sodium sulfate. The benzene solution is stirred with rnercury overnight, then centrifuged and the clear benzene solution is evaporated in vacuo. The residual material thus obtained is separated using partition chromatography on diatomaceous silica (Supercel) to give llB,l7a,-21 trihydroxy-6a,l6m-dimethyl-1,4-pregna diene-3,20-dione ZI-acctate. hydroxy-6a,'16a-dimethyl-l,4-pregnadiene-3,ZO-dione 2lacetate is dissolved in a mixture of 1.0 cc. of benzene and 1.0 cc. of 1 N methanolic potassium hydroxide, and the solution is allowed to stand at room temperature for a period of about 10 minutes. The solution is then acidified with acetic acid, then benzene is evaporated in vacuo and the residual material is purified by crystallization to give ll5,l7a,2l-trihydroxy-6a,16o-dimethyl-1,4-pregnadiene-3 ,20-dione.

' Similarly, in accordance With the foregoing procedures but starting with 1l5,l7u,2l-trihydroxy-6a,ldB-dimethyl- 4-pregnene-3,20-dione 2l-acetate, there are obtained the corresponding 11 8, l7a-21-trihydroxy 6oc,16,8 dimethyll,4-pregnadiene-3,20-dione, and its Ill-acetate.

To a cooled solution of 436 mg. of llfi,l7a,21-trihy- 2l-acetate (which can be prepared as described hereinabove) in 2.5 cc. dimethyl formamide and 2.0 ml. of dry pyridine is added 1.0 ml. of methane sulfonyl chloride, while maintaining the temperature below 0 C. The resulting mix ture is allowed'to warm to room temperature, at which heated to a temperature of about 70-l00 C. for a period of about 10 minutes. About 15 ml. of water is added slowly to the reaction mixture, with stirring, and the aqueous mixture is extracted with ethyl acetate. The combined ethyl acetate extracts are washed with water, then with dilute aqueous hydrochloric acid solution, again with water, and then with a dilute aqueous sodium bicarbonate solution. The washed ethyl acetate solution is dried, and the solvent is evaporated in vacuo; the residual material is triturated with ether, and the crystalline material is recrystallized to give 17,2l-dihydroxy-6a,l6cx-dimethyl-4,9 (ll)-pregnadiene-3,20-dione 21-acetate. A suspension of 330 mg. of 17a,21-dihydroxy-6u,l6zx-dimethyl-4,9(1l}- pregnadiene-3,20-dione 2l-acetate and 1.8 g. of N-bromosuccinimide in a mixture of 50 ml. of dioxane and 10 ml. of water is cooled to 10 C. Then with stirring, 10 ml. of cold 1.0 N aqueous perchloric acid is added. The temperature of the reaction mixture is allowed to rise to 15 C. and is maintained at this point for about two and one-half hours during which time the solid material slowly dissolves. The resulting yellow solution is treated with 1.0 ml. of allyl alcohol to discharge the color and the remaining N-bromo-succinimide, and the resulting solution is evaporated to a small volume in vacuo. The concentrated solution is diluted with water, and the aqueous mixture is extracted with ethyl acetate. The ethyl acetate extracts are washed, dried and evaporated to dryness, and the residual material is crystallized from ethyl acetateether to give 9a-bromo-11B,1Za.21-trihydroxy-6a,16a-dimethyl-4-pregnene-3,ZO-dione ZI-acetate.

A solution of 210 mg. of 9ot-bromo-1lfii,l7u,2l-trihydroxy-6a,16a-dimethyl-4-pregnene-3,ZO-dione Zl-acetate and 240 mg. of potassium acetate in 10 ml. of absolute ethanol is heated under reflux for two hours. The reaction mixture is cooled to room temperature, evaporated in vacuo to a small volume, and diluted with water. The concentrated aqueous mixture is extracted with three portions of ethyl acetate, and the combined ethyl acetate extracts are washed with water, dried, and evaporated to dryness in vacuo. The residual material is crystallized from ethyl acetate-ether to give 95,115-6POXY-17oa21-dihydroxy-6or,l6oz-dimethyl-4-pregnene-3,20-dione 2l-acetate.

To a solution of 83 mg. of anhydrous hydrogen fluoride in 4.7 ml. of ice-cold alcohol-free chloroform is added an ice-cold solution of 416 mg. of 9,3,11fi-POXy-170L,21'dlhydroxy-6a,16a-dimethyl-4-pregnene-3,ZO-dione 21-acetate. The solution is mixed thoroughly and maintained at C. for two hours, at the end of which time 13.0 ml. of ice-cold 20% aqueous sodium acetate is added, and the resulting mixture agitated vigorously. The layers are separated, and the chloroform layer is washed with water until free of acid, dried, and the chloroform evaporated in vacuo. The residual material is crystallized from acetone-petroleum ether to give 9a-fluoro-11B,17u,21-trihydroxy-6cx,l6a-dimethyl-4-pregnene-3,ZO-dione 2l-acetate. Fifty milligrams of 9or-fiuoro-1lfi,l7a,2l-trihydroxy-6a, 16or-dimethyl-4-pregnene-3,20-dione ZI-acetate is dissolved in a mixture of 1.0 cc. of benzene and 1.0 cc. of 1 N methanolic potassium hydroxide, and the solution is allowed to stand at room temperature for a period of about 10 minutes. The solution is then acidified with acetic acid, the benzene is evaporated in vacuo, and the residual material is purified by crystallization to give 9a-fiuoro-115, 17a,2l-trihydroxy-tia,l6a-dimethyl-3,20-dione.

Similarly, when hydrogen chloride is substituted fo hydrogen fluoride in the reaction with 9,8,llfl-epoxy-17a, 2l-dihydroxy-6ot,16oz-dimethyl-4-pregnene-3,20-dione 21- acetate there is obtained the corresponding 9os-Ci1l0r0-1lfl, 17 x-21-trihydroxy-6et,16ct-dimethyl-4-pregnene-3,ZO-dione.

In accordance with the foregoing procedures when 11 8, 17,21 t'rihydroxy-oa,16;?-dimethyl-4-pregnene-3,20-dione 21-acetate is employed as the starting material, there is otbained' the corresponding 9ix-fiuoro-1lfi,l7ot,2l-trihydroxy-6a,16,B-dimethyl 4 pregnene-3,20-dione or 9a- 28 chloro-l1,8,17a,2l-trihydroxy-6 x,16,8-dimethyl-4 pregnene- 3,20-dione.

To a solution of 100 mg. of 9ot-iiuoro-11B,17a., Zl-trihydroxy-6ot,l6a-dimethyl-4-pregnene-3,ZO-dione 21-acefate, in 6 ml; t-butanol, and 0.01 ml. of glacial acetic acid is added 70 mg. of selenium dioxide. The mixture is heated under reflux in a nitrogen atmosphere for a period of about 60 hours. The reaction solution is filtered and evaporated in vacuo. The residual material is dissolved in benzene, and the benzene solution is washed with an aqueous sodium bicarbonate solution and dried over anhydrous sodium sulfate. The benzene solution is stirred with mercury over-night, then centrifuged, and the clear benzene solution is evaporated in vacuo. The residual material thus obtained is separated, using partition chromatography on diatomaceous silica (Supercel), to give 9a fluoro 1lB,l7a,21 trihydroxy-a,l6adimethyl-1,4- pregnadiene-3,20-dione 21-acetate.

This material is hydrolyzed using 1 N methanolic potassium hydroxide in benzene as described above for the hydrolysis of 9cx-fiuoro-11B,17a,21-trihydroxy-6a,16a-dimethyl-4-pregnene-3,ZO-dione 21-acetate to give 9a-fiuoro- Y 11B,17ot,2l-trihydroxy-6a,16a-dimethyl-1,4 pregnadiene- 3,20-dione.

Starting with t-ChlOlO-11fi,17a,21-l1lhydlOXy-6zx,160tdimethyl-4-pregnene-3,20-dione 21-acetate, there are obtained in accordance with the foregoing procedures the corresponding 9a-chloro-11B,l7ot,21-trihydroxy-6u,l6ct-d1- methyl-l,4-pregnadiene-3,ZO-dione, and its Zl-acetate.

Similarly, starting with 9a-brorno-l1fi,l7a,21-trihydroxy-6a,16u-dimethyl-4-pregnene3,20-dione ZI-acetate obtained as an intermediate in the preparation of 90:- fluoro 11{3,l7a,21 trihydroxy-6a,16a-dimethyl-4-pregnene-3,20-dione herein described above, there are obtained the corresponding 9ot-bromo-1 1 [8,170t,21-t1'lhydYOX 6a,l6a-dimethyl 1,4-pregnadiene-3,ZO-dione, and its 2l acetate. 7

In accordance with the foregoing procedures, but start ing with 9rx-halo-ll,8,17u,21-trihydroxy-6ot,16fi-dimethyl 4-pregnene-3,20-dione, 21-acetate there are obtained the corresponding 9ot-halo-l1B,l7u,21-trihydroxy-6u,16B-dimethyl-1,4-pregnadiene-3,20-diones such as for example 911 fluoro 11B,170,21-trihydroxy-6a,lfifl-dimethyl-1,4- pregnadiene-3,20-dione, 9ct-chloro-l1B,17oc,21trihydroxy- 6a.,16B-dimethyl-l,4-pregnadiene-3,20-dione and 90c-b10- mo-l1B,17a,2l-trihydroxy-6a,l6,B dimethyl 1,4-pregnadiene-3,20-dione, and their 21-acetates.

A solution of 400 mg. of 9u-fiuoro-11[3,l7a,21-trihydroxy-6a,16a-dimethyl 4 pregnene-3,20-dione Zl-acetate in 4 ml. of pyridine is added to the complex formed by the addition of 400 mg. of chromium trioxide to a 4 ml. of pyridine. The mixture is swirled until thoroughly mixed and then allowed to stand at room temperature overnight. The reaction mixture is poured into water, and the aqueous mixture is extracted with ether and then twice with ethyl acetate. The combined ether and ethyl acetate extracts are washed with dilute aqueous sulfuric acid at about 0 C., and then with water until neutral. The organic solvent layer is then dried, the solvents are evaporated therefrom in, vacuo, and the residual crystalline material is purified by crystallization to give 9a-fluoro-17ot,21-dihydroxy-6ot,l6tx-dimethyl-4- pregnene-3,l1,20-trione ZI-acetate. Fifty milligrams of 9a-tluoro-17u,2l-dihydroxy 60.,l6ix dimethyl-4-pre nene-3,1l,20-trione 21-acetate is dissolved in a mixture of 1.0 cc. of benzene and 1.0 cc. of 1 N methanolic potassium hydroxide, and the solution is allowed to stand at room temperature for a period of about 10 minutes. The solution is then acidified with acetic acid, the benzene is evaporated invacuo, and the residual material is purified by crystallization to give 9o:-flUOIO-17ot,2l-dihydroxy-6a,16ot-dimethyl-4-pregnene-3 ,1 1,2 O-trione.

Similarly, starting with 9a-chloro-11,8,17a,21-trihydrOXy-6oc,l6oc dimethyl-4-pregnene-3,20-dione 21-acetate, there are obtained in accordancewiththe foregoingprotate.

V gen. acetate and filtered. The filtrate is Washed sequentially 2 cedures the corresponding 9a-chloro- 17a,2l-dihydroxy- 6oz,16a-dimethyl-4-pregnene-3,11,20-trione, and its 21-a ce tate. a w N f Similarly, but starting with 9a-bronio-ll 3,l7a,2l-tri hYdIOXY-Goc,16ct-dilll6ih3/l-4-Pl6gll8l'l6-3,ZO dlQIIe 2 l-acetate obtained as an intermediate in the preparation of 9afluoro -;11;3,17a,21 trihydroxy-6ot,16d9dimethyl-4-pregphase isdried over sodium sulfate and concentrated in vacuo. I The crude material dissolved in benzene 1s chro-' matographed on 15 g. of acid-washed alumina. Elution with chloroformfether 2:8 atiords 17u,21-dihydroxy- 6,16a-dimethyl-4,6-pre'gnadiene-3 ,1 1,20-trione 21-acetate.

- tion of potassium bicarbonate in aqueous methanol to nene-3,2 dione herein described above, ther'e are ab 7 tained "the correspondingfl 9ct-bromo-l7iii,21-dihydroxy-jl 6a,16st-dimethyl-4-pregnene-3,11,%0-trione,"and its"2l :ace-

' In accordance with the-foregoing procedures, istart- 4-pregnene-3,20-dione 21-acet'ate, there are'obtained the corresponding 9a-halo-17a,21-dihydroxy-6 x,l6f3-dimethyl- "'4rpregnene-3,11,20 triones,. such as for example, 9%

droxy-6u,16asdimethyl-4-pregnene-3,11,20 trione 21-acei. In accordance wrth lthe above procedure but starting In accordance with the above procedure but starting with; 1 15,17 11,21-1trihydroxy-6a,16a-dirnethyl-4-pregnene- 3,20dione 21-acetate there is obtained '11 8,17a,21-'trihydroxy-6,16a-dimethyl 4,6-pregnadiene-3,20 dione and tate, in 6 ml. t-butanol, and 0.01 ml.,of glacial acetic acid is'added 70 mg. of selenium dioxide. The mixture is its Zl-acetate.

In accordance with the above procedure'but starting with 11B,17a,2l-trihydroxy-6a,16,8-dimethyl-4-pregnene- 3,20-dione 21-acetate there is obtained 11fi,l7ot,21-trihydroxy-G,165-dirnethyl-4,6-pregnadiene-3,20 dione and its '2l-acetate. a

A suspension containing 300 mg. of 9ot-fill01O-11fi,17oc,

ofk about 60 hours. jThe reaction solution is filtered" andevaporated in vacuo. The residual material is dis- 7 solved in benzene, and the benzene solution is washed with-an aqueous sodiur'n'bicarbonate solution and dried over anhydrous sodium sulfate. Thebenzene solution :is stirred with mercury overnight, then centrifuged, and

the clear benzene solution is evaporated in vacuo.

The residual material thus obtained is separated using partition chromatography on diatomaceous silica (Supercel) to give 9a-fluoro-17a,2.1-dihydroxy-6rx,16u-dirnethyl-l,4- pregnadiene-3,11,20-trione 2l-acetate. This material is hydrolyzed using 1 N methanoli c potassium hydroxide in benzene as described hereinabove for the hydrolysis of 9a-fiuoro l7oc,21 dihydroxy-6ot,l6e-dimethyl-4-pregnene-3,l1,20-trione 2l-acetafe .to give 90t-fiI1OIO-170t,21- dihydroxy 6a,16a dimethyl-1,4-pregnadiene-3,1l,20-tri one.

Similarly, starting with 9ot-chloro-17a,21-dihydr0xy- 6ogl6a-dimethyl 4 pregnene-3,ll,20-trione 21-acetate there are obtained in accordance with the foregoing procedures the corresponding 9a-chloro -17o,2l-dihydroxy- 6a,lmsdimethyl-l,4-pregnadiene-3,11',20:trione and. its 2l-acetate.

Similarly, but starting with 9a-bromo-l7a,21-dihydroxy- 60,160t-dii'l'l6thYl- 4 -pregnene-3,l1,20-trione 2l-acetate there are obtained the corresponding'9a-bromo-l7 2l-dihydroxy-6a,l6a-dimethyl-1,4-pregnadiene 3,11,20-trione,

6a,l6wdimethyl-4-pregnene-3,11,20-trione Ill-acetate and 1.7 ml. of acetic acid is refluxed for 17' hours under nitro- The reaction mixture is diluted with 40 ml. of ethyl with two 20 ml. portions of ice cold 10% aqueous sodium bisulfite solution, three 20 ml. portions of ice cold 5% aqueous potassium hydroxide solution and finally Washed to neutrality with several 20 ml. portions of ice water.

.;The aqueous washes are re-extracted with mlof'ethyl acetate and treated as above. The combined organic acetate, 700mg. of chloranil, 8.3 ml. of ethyl acetate and 1.7 ml. of acetic acid is refluxed for 17 hours under nitroacetate and filtered. The filtrate is washed sequentially with two 20 ml. portions of ice cold 10% aqueous sodium bi sulfite solution,'three 20 ml. portions of ice cold 5% aqueous potassium hydroxide solution and finally washed to neutrality with several 20 ml. portions of ice water. The aqueous washes are re-ex-tracted with 50 ml. of ethyl acetate and treated as above. The combined organic phase is dried over sodium sulfate and concentrated in vacuo. The crude material dissolved in benzene is chro: matographed on 15 g. of acid-washed alumina. Elution with chloroformzether 2:8 afiords 9'at-flll010-1lfi,17oc,21 trihydroxy 6,160: dimethyl 4,6 pregnadiene 3,20- dione 21-acetate. This product is then hydrolyzed by treatment with a solution of potassium bicarbonate in aqueous methanol to form 9a-fluoro-11fi,17u,21trihydroxy-6,16u-dimethyl-4,6-pregnadione-3,20-dione.

In accordance with the above procedure but starting with 9ot-chloro-l 1,8,17a,21-trihydroxy-6a,16a-dimethyl-4- pregnene 3,2G-dione 2l-acetate-, there are obtained the corresponding 9,0t-Cl1l01O-1 1B,17ot,21-trihydroxy-6,l6u-dimethyl-4,6-pregnadiene-3,20-dione and'its ZI-acetate.

In accordance with the above procedure but starting with 9ot-bromo-11B,l7o,21-trihydroxy-6,16a-dimethyl-4- pregnene-3,20-dione ZI-acetate, there are obtained the corresponding 9ot e bromo 11fi,17oc,21 trihydroxy-6,16a-diinethyl- 4-,6-pregnadiene-3,ZO-dione and its 2l-acetate.

Similarly, in accordance with the above procedure but starting with 9a-fiuoro-11,8,17a,21-trihydroxy-6a,16B-dimethyl-4-pregnene-3,ZO-dione ll-acetate, there is obtained the corresponding 9a-fluoro-llfi,l7a,21-trihydroXy-6,165- dimethyl-4,6-pregnadiene-3,20-dione and its 21-acetate.

Similarly, in accordance with the above procedure but starting with 9ot-chlororll5,17a,21trihydroxy-6a,l6B-dimethyl-4 -pregnener3,ZO-dione Zl-acetate, there is obtained the corresponding 9ot-chloro-11,6,l7a,2l-trihydroxy-6,16fldimethyl-4,6-pregnadiene-3,20-dione and its 21-acetate. Similarly, in accordance with the above procedure but starting with 9ot-brorno-11B,lfla,2l-trihydroxy-6a,16,8-dimethyl-4-pregnene-3,ZO-dione 21-acetate, there is obtained the corresponding 9a-bromo-1 1B, 171:,2l-trihydroxy-6,16;3-'

,dimethyltfi-pregnadiene-3,ZO-dione and its 21-acetate.

A suspension containing 300 mg. of 9oz-fiuoro-17a,21-

dihydroxy-6a,l6ot-dimethyl-4 pregnene-3 ,1 l,20-trione 21 acetate, 700 mg. of chloranil, 8.3 m1. of ethyl acetate and mLof acetic acid. is refluxed for 17 hours under nitrogen. The reaction mixture is diluted with 40 ml. of ethyl acetate and filtered. The filtrate is washed sequentially with two 20 ml. portions of ice cold aqueous sodium bisulfite solution, three 20 ml. portions of ice cold 5% aqueous potassium hydroxide solution and finally washed to neutrality with several 20 ml. portions of ice Water. The aqueous washes are re-extracted with 50 ml. of ethyl acetate and treated as above. The combined organic phase is dried over sodium sulfate and concentrated in vacuo. The crude material dissolved in benzene is chrowith 9a-chloro-17a,21-dihydroXy-6ot,l6tx-dimethyl-4-pregnene-3,1l,20-trione ZI-acetate there is obtained the corresponding 9a-chloro 17a,21-dihydroxy-6, 16ot-dirnethyl-4,6- pregnadiene-3,11,20-trione and its 2l-acetate.

In accordance with the above procedure but starting with 9a-bromo-l7a,21-dihydroxy-6 x,16a-dimethyl-4-pregnene-3,11,20-trione ZI-acetate there is obtained the corresponding 9 c -bron1o-17o;,2l-dil1ydroxy-6,16a-dimethy1-4,6- pregnadiene-Zad1,20-trione and its ZI-acetate.

Similarly, in accordance with the above procedure but starting with 9ot-fiuoro- 17a,21-dihydroxy-6ot,lfi-dirncthyl- 4-pregnene-3,-11,20-trione ZI-acetate, there are obtained the corresponding 9a-fluoro-17a,2l-dihydroxy-6,16,B-dimethyl-4,6-pregnadiene-3,11,20-trione and its ZI-acetate.

Similarly, in accordance with the above procedure but starting with 9oc-ch1oro-17a,21-dihydroxy-6ot,l6/3-dimethyl- 4-pregnene-3,11,20-trione ZI-acetate, there are obtained the corresponding 9ot-chloro-17a,2l-dihydroxy-6,l6fl-dimethyl-4,6-pregnadiene-3,11,20-trione and its 21-acetate.

Similarly, in accordancewith the above procedure but starting with 9tx-bromol7 x,2l-dihydroxy-6u,IGB-dimethyl-4-pregnene-3,11,20-trione ZI-acetate, there are obtained the corresponding 9a-bromo-17a,21-dihydroxy-6,16fl-dimethyl-4,6-pregnadiene-3,11,20-trione and its 2l-acetate.

To a solution of 200 mg. of 17u,2l-dihydroxy-6,l6u dimethyl-4,6-pregnadiene-3,11,20-trione 2l-acetate in 0.16 ml. of acetic acid and 12 ml. of t-amyl alcohol is added 120 mg. of selenium dioxide and 2 drops of mercury. The mixture is refluxed under nitrogen overnight. The solution is'filtered, washed with sodium bicarbonate, dried, and concentrated. This product is chromatographed on acid-washed alumina. The adsorbate is eluted with mixtures of chloroform and ether, increasingly rich in chloroform. The eluates are combined, evaporated to dryness and the residual material recrystallized from ethyl acetate to give l7a,21-dihydroxy-6,16a-dimethyl-1,4,6-pregnatrione-3,11,20-trione 2l-acetate. This product is then'hydrolyzed by treatment with a solution of potassium bicarbonate in aqueous methanol to form l7ot,2l-dihydroxy- 6,16a-dimethyl-1,4,6-pregnatriene 3 ,11,20-trione.

In accordance with the above procedure but starting with 17a,2l dihydroxy-6,16,8-dimethyl-4,G-pregnadiene- 3,11,20-trione 2l-acetate there are obtained l7a,2l-dihydroxy 6,165 dimethyl-1,4,6pregnatriene-3J1,20-trione and its ill-acetate.

Similarly, when 11fl,l7a,21-trihydroxy-6,16u-dimethyl- 4,6pregnadiene-3,20-dione 2l-acetate is used as starting material in the foregoing reaction with selenium dioxide and the product hydrolyzed with aqueous methanolic potassium bicarbonate solution, there are obtained 1113,17a, 21 trihydroxy 6,16a-dimethyl-1,4,6-pregnatriene-3,20-

vdione and its 21-acetate. Where the starting material in this procedure is 1l/3,l7o,2l-trihydroXy-6,16,8dirriethyl-4, 6-pregnadiene-3,ZO-dione Zl-acetate, thereare obtained 7 1 trihydroxy 6,16ot-dimethyl-1,4,6-pregnatrieneone-3,20-dione and its 21-acetate.

Where the starting material in the foregoing reaction 7 with sel n um dioxide followed by hydrolysis with aqueous methanolic potassium bicarbonate is 9u-fiuoro-llfl,

1704,21 tri'nydroxy 6,l6ot-dimethyl-4,6-pregnadiene-3,20- dione 2l-acetate, the products obtained are 9a-tluoro-l1fl, l7ot,2l 'trihydroxy 6,l6a-dimethyl-1,4,6-pregnatriene- 3,20dione and its ZI-acetate.

, In accordance with the above procedure but starting with 9a-chloro-1 176,17x,2lrtrihydroxy-o,l6ot-din1ethyl-4,6- pregnadiene-3,20-dione 2l-acetate there are obtained 9a- ChlOIOf'lL ,17OL,2l.-tfihYdfOXY-6,1 60C'CllITlthy1-1,4,6 pregnadiene-3,20-dione and its 2l-acetate. 5 I

Similarly, inaccordance with the above procedure but starting with 9a-bromoel 1 fl,l7u,2 l-trihydroxy-o, 160t-dlmethyl-4,6-pregnadiene-3,ZO-dione 21-acetate there are obtained 9otbromo- 11,8,17ot,21-trihydroxy-6,l6a-dimethyl-1,4,6-pregnatriene-3,20-dione and its Ill-acetate.

In accordance with the above procedure but starting with 9ot-fluoro-l1 fi,170t,21-lllll1ydfOX -6,l6fi-dimethyl-4,6-

pregnadiene-3,20-dione ZI-acetate there are obtained 9afiuoro 1 16,17a,21-trihydroxy-6,16B-dimethyl-l,4,6-pregnatriene-3,20-dione and its 2l-acetate.

In accordance with the above procedure but starting with 9a-chloro-l1,8,17a,2l-trihydroxy-6,16,3-dimethyl-4,6- pregnadiene-BQU-dione 21-acetate there are obtained 90.- chloro-1lB,l7a,21-trihydroXy-6,l 6fi-dimethyl-l,4,6, pregnatriene-3,20-dione and its 21-acetate. g

. In accordance with the above procedure but starting pregnadiene-3,20-dione 2l-acetate there are obtained bromo-l1p,l7a,21-trihydroXy-6,l6B-dimethyl-1,4,6 pregnatriene-3,20-dione and its 2l-acetate. f

i To a solution of 200 mg. of 9a-duoro-l7m2l-dihydroxy-'6,1'6a-dimethyl-4,6-pregnadiene-3,1 1,20-trione 21- acetate in 0.16 ml. of acetic acid and 12 ml. of t-amyl alcohol is added mg. of selenium dioxide and 2 drops of mercury. The mixture is refluxed under nitrogen overnight. The solution is filtered, washed with sodium bicarbonate, dried, and concentrated. This product is chromatographed on acid-Washed alumina. The-adsorbate is eluted with mixtures of chloroform and ether, increasingly rich in chloroform. The eluates are combined, evaporated to dryness and the residual material recrystallized from ethyl acetate to give 9ocfillOI'O-17zx,21 dihydroxy 6,l6u-dirnethyl-1,4,6-pregnatriene-3,11,20-trione 2l-acetate. This product is then hydrolyzed by treatment with a solution of potassium bicarbonate in aqueous methanol to form 9oa-fluoro-17ot,2l-dihydroXy-6,l6tx-dimethyl- 1,4,6-pregnatriene-3,1 1,20-trione.

In accordance with the above procedure but starting with 9a-chl0ro-17a,21-dihydroxy-6,l6a-dimethyl-4,6-pregnadiene-3,11,20-trione 21-acetate there are obtained 9achloro 17a,21-dihydroxy-6,16a-dimethyl-l,4,6 pregnatriene=3,l1,20-trione and its ZI-acetate.

Similarly, in accordance with the above procedure but starting with 9a-bromo-l7a,2l-dihydroxy-6,16ot-dimethyl- 4,6-pregnadiene-3,11,20-trione 21-acetate there are obtained 9a-bromo-l7a,21-dihydroxy-6,16a-dimethyl-1,4,6- pregnatriene-3,11,20-trione and its 21-acetate.

In accordance with the above procedure but starting with 9a-fluoro-17a,2l-dihydroxy-6,16fi-dimethyl-4,6-pregnadiene-B ,ll,20-trione 21-acetate there are obtained 9afiuoro :,21 dihydroXy-6,16,8-dimethyl-1,4,6-pregnatriene-3,11,20-trione and its 21-acetate.

In accordance with the above procedure but starting with 90L-ChlOIO-17m,21dlhydI'OXy'6, 1 6fi-dirnethyl-4,6-pregnadiene-3,l1,20-trione ZI-acetate there are obtained 90:-

chloro l7ot,21-dihydroXy-6,16B-dimethyl-1,4,6 pregna- 33 l. of alcohol-free chloroform cooled to about C. in an ice bath is added with constant stirring 750 ml. of cold, concentrated aqueous hydrochloric acid solution and then 740 ml. of formalin (low in methanol). The mixture is removed from the ice bath and stirred at room temperature for approximately 7 hours. The layers are separated and the aqueous phase is back-extracted twice with chloroform. The combined chloroform layers are Washed twice with a 5% aqueous solution of sodium bicarbonate, and twice with a saturated aqueous solution of sodium chloride. The washed chloroform solution is dried over magnesium sulfate and evaporated under reduced pressure. The residual material is triturated with methanol to give a crystalline solid, which is recrystallized from a mixture of benzene and. n-hexane to give 17u,20,20,21-bismethylenedioxy-11,8-hydroxy 6, 16a dimethyl- 4,6-pregnadiene-3-one.

A solution of 400 mg. of 17a,20,20,2l-bismethylenedioxy 115 hydroxy-6,16wdimethyl-4,6 pregnadiene-3- one in 4 ml. of pyridine is added to the complex formed bythe addition of 400mg. of chromium trioxide to 4 ml. of pyridine. The mixture is swirled until thoroughly mixed and then allowed to stand at room temperature for a period of about 15 hours. The reaction mixture is poured into Water, and the aqueous mixture is extracted with ether and then twice with ethyl acetate. The combined ether. and ethyl acetate extracts are washed with dilute aqueous sulfuric acid at about 0 C., and then with water until neutral. The organic solvent layer is then dried, the solvents are evaporated therefrom in vacuo,

and the residual crystalline material is purified by crystallization from a mixture of ethyl acetate and ether to give 17u,20,20,2 l-bismethylenedioxys6,16a dimethyl-4,6- pregnadiene-lllsdione.

Approximately 1.35 g. of 17a,20,20,2l-bismethylene- ,dioxy 6,161x. dimethyl 4,6 pregnadiene 3,11-dione is dissolved in 23 ml. of dry, hot benzene, the resulting solution .is cooled to room temperature, and to the solution is added 0.96 ml. of freshly-distilled ethyl formate. The air in thesystem is-replaced with nitrogen and 560 mg.- of sodium hydride (as a 58% dispersion in mineral oil) is added. The system is again evacuated and filled with nitrogen, and the mixture is stirred at room temperature for a period of about 15 hours. The reaction mixture isv poured into an'excess of a saturated aqueo u g solution of sodium dihydrogen phosphate, and the resulting aqueous mixture is extracted four times with benzene. The benzene reaction solution and extracts are combined, washed three times with Water, dried over sodium sulfate, and evaporated to dryness. The residual material is dissolved in ether, and the ether solution is extracted with a aqueous solution of sodium carbonate. The aqueous alkaline extracts are acidified with an excess of a saturated aqueous solution of sodium dihydrogen phosphate, and the aqueous acidic mixture extracted with ether and then with chloroform. The combined organic extracts are dried over sodium sulfate, and evaporated to dryness to give 170;,20,20,21-bismethylenedioxy-2 hydroxymethylene 6,160 dimethyl 4,6 pregnadiene- 3, 1'1-dione.

Approximately 850 mg. of l7u,20,20,2l-bismethylene-,

dioxy 2 hydroxymethylene 6,160 dimethyl 4,6-

pregnadiene-Lll-dione is dissolved in 9.2 ml. of absolute ethanol, and to the solution is added a solution of 0.16 ml. of hydrazine hydrate dissolved in 0.16 ml. of absolute ethanol. The resulting mixture is heated under reflux in 7 a nitrogen atmosphere fora'period of about 45 minutes,

and the reaction mixture is thenevaporated to dryness under reduced pressure. The residual material is washed with three portions of cold water, and the'resulting amordioxy 6,16ot dimethyl [3,2-c1pyrazolo 4,6 pregnadiene-ll-one in 2 ml. of pyridine is added 0.5 ml. of acetic anhydride. The mixture is allowed to stand overnight at room temperature. A mixture of ice and water is then added. After standing for about 30 minutes, the product is extracted with ethyl acetate. The ethyl acetate extract is washed successively with water, with ice-cold 1 N sulfuric acid (until the pH ofthe aqueous layer is 1-3), (with saturated aqueous sodium bicarbonate solution (until the pH of the aqueous layer is 8), and with water (until the aqueous layer is neutral). The ethyl acetate solution is then dried with anhydrous sodium sulfate, and the solvent is distilled at a temperature of about 40 C., in vacuoto give 1-acetyl-17a,20,20,21-bismethylenedioxy 6,16a dimethyl [3,2-c] pyrazolo 4,6 pregnadiene-ll-one, which is isolated by the addition of water and filtration.

In accordance with the above procedure, but using an equivalent quantity of another acyla-ting agent in place of the acetic anhydride, there is obtained the corresponding l'-acyl-derivative.

Approximately 720mg. of 1'-acetyl-17a,20,20-21-bismethy-lenedioxy 6,=l6 x dimethyl [3,2-c]pyr-azolo 4,6- pregn-adiene-ll-one is heated on a steam bath with 24 ml. of a 60% aqueous solution of formic acid for a period of about 30 minutes. The excess reagent is removed in vacuo using a Water bath at a temperature of about 50 C. as the source of heat. The residual material is flushed four times with n-hexane, and dried at 60 C. in high vacuum. The amorphous solid thus obtained is dissolved in about 3.4 ml. of pure methanol and allowed to react with 1.3 ml. of a 1.33 N solution of sodium methoxide in methanol at room temperature under a nitrogen atmosphere for ten minutes. The excess sodium methoxide is neutralized with acetic acid, and the mixture is then evaporated to dryness and flushed with n hexane. The residual material is washed with water, filtered and dried to give 17u,21-dihydroxy-6,Mot-dimethyl-[3,2-c1pyrazolo- 4,6-pregnadiene-1-1,20-dione.

In accordance with the above procedures, but starting with the 1lB,-17a,2l-trihydroxy-6,16fi-dimethyl-4,6-pregnadiene-3,20-dione (which can be prepared as described in Example 20 hereinabove) in place of the 11B;17oc,21 trihydroxy 6,160; dimethyl 4,6 4 pregnadiene 3,20- dione, there is obtained as product the corresponding 1711,21 dihydroxy 6,1618 dimethyl [3,2-c]pyrazolo- 4, 6-pregnadiene-1 1,20-dio-ne.

In accordance with the above procedures, but starting with 11fi,17rx,2)1 trihydroxy 6,160: dimethyl 9afluoro-4,6-pregnadiene-3,20-dione (which can 'be prepared 53 described in Example 28 hereinabove) in place of the :lfi,17a,2l trihydroxy 6,16a dimethyl 4,6 pregna- Jiene-3,20-dione, there is obtained as product, via the intermediate formation of 1'-acetyl-17a,20,20,2l-bismethy-lenedioxy 6,1611 dimethyl 9a fluoro [3,2-c]pyrazolo-4,6-pregnadiene-1l-one, the corresponding 17a,21-di hydroxy 6,16a dimethyl 9a fiuoro [3,2-c1pyrazolo- 4,6-pregnadiene-l1,ZO-dione. Alternatively, the intermediate 1' acetyl 17a,20,20,2l bismet-hylenedioxy 6, 16 dimethyl 9a fluoro [3,2-c1-pyrazolo 4,6 pregnadiene-ll-one is reacted with sodium borohydride to [forml acetyl 17a,20,20,21 bismethylenedioxy 6, 16oz. dimethyl 9oz flu-oro [3,2-c1pyrazolo 4,6 pregnadiene-llB-ol which is then reacted with 60% aqueous tormic acid and the resulting amorphous solid reacted with dilute methanolic sodium methoxide as described hereinabove in this example, thereby forming 11/3,17a, '21 trihydroxy 6,16u dimethyl a fluoro [3,2-c]- pyrazolo-4,6-pregnadiene-20-one.

Approximately 1.35 g. of 17o,20,20,21 bismethylenedioxy 11B hydroxy 6,160: dimethyl 4,6 pregnadiene-3-one (which can be prepared as described hereinabove in this example) is dissolved in 23 'ml. of dry, hot benzene, the resulting solution is cooled to room temperature, and to the solution is added 0.96 ml. of freshlynight at room temperature.

product is extracted With ethyl acetate. extract is Washed successively with water, with ice-cold about '30 minutes.

' 35 distilled ethyl formate. The air in the system is replaced with nitrogen and 560 mg, of sodium hydride (as a 58% dispersion in mineral oil) is added. The system is again evacuated and filled with nitrogen, and the mixture is stirred at room temperature for a period of about hours. The mixture is poured into an excess of a saturated aqueous solution of sodium dihydrogen phosphate, and the resulting aqueous mixture is extracted four times with benzene. The benzene reaction solution and extracts are combined, washed three times with water, dried over sodiumv sulfate, and evaporated to dryness. The residual material is dissolved in ether, and the ether solution is extracted with a 10% aqueous solution of sodium carbonate. The aqueous alkaline extracts are acidified with an excess of a saturated aqueous solution of sodium dihydrogen phosphate, and the aqueous acidic mixture extracted with other and then with chloroform. The combined organic extracts are dried over sodium sulfate, and evaporated to dryness to give 17oc,20,20-2l-bi8m6thylenedioxy 11p hydroxy 2 hydroxymethylene 6, 16a-dimethyl-4,6-pregnadiene-3-one.

Approximately 850 mg. of 17a,20,20,2l-bismethylenedioxy 11,8 hydroxy 2 hydroxymethylene 6,16adimethyl-4,6-pregnadiene-3-one is dissolved in 9.2 ml. of absolute ethanol, and to the solution is added a solution of 0.16 ml; of hydrazine hydrate dissolved in 0.16 ml. of absolute ethanol. The resulting mixture is heated under reflux in a nitrogen atmosphere tor a period of about 45 minutes, and the reaction mixture is then evaporated to dryness under reduced pressure. The residual material is washed with three portions of cold water, and the resulting amorphous solid is dried at atemperature of about 80 C. for 1 hour in high vacuumto give 170L,20,20,21- bismethylenedioxy 6,1604 dimethyl' -'[3,2-c]pyrazolo- 4,6-pregnadiene-l lfl-ol.

To a solution of 100 mg. of 17a,20,20,2l bismethylenedioxy 6,160: dimethyl [3,2-c]pyrazolo 4,6 pregnadiene-llB-ol in 2 ml. of pyridine is added 0.5 ml. of acetic anhydride. The mixture is allowed to stand over- A mixture of ice and water is then added. After standing for about minutes, the The ethyl acetate 1 N sulfuric acid (until the pH of the aqueous layer is -13 with saturated aqueous sodium bicarbonate solution (until the pH of the aqueous layer is 8), and with water (until the aqueous layer is neutral). The ethyl acetate solution is then dried with anhydrous sodium sulfate, and the solvent is distilled at a temperature of about C.

in vacuo to give 1'-acetyl-17u,20,20,2l-bismethylenedioxy 6,16u dimethyl [3,2-.c]pyrazolo 4,6 pregnadiene-llfi-ol which is isolated by the addition of water and filtration.

In accordance with the above procedure, but using an equivalent quantity of another acylating agent in place of acetic anhydride there is obtained the corresponding 1'- .acyl-steroid.

Approximately 720 mg. of l-acetyl-l7a,20,20,2l-bispregnadiene-llB-ol is heated on a steam bath with 24 m1. of a 60% aqueous solution of formic acid for a period of The excess reagent is removed in vacuo using a Water bath at a temperature of about 50 C. as the source of heat. The residual material is flushed four times [with n-hexane, and dried at 60 .C, in high vacuum. The-amorphous solid thus obtained is dissolved in .about 3.4 ml. of pure methanol and allowed to react with 1.3 ml. of a 1.33 N solution of sodium methoxide in methanol at room temperature under a nitrogen atmosphere .for ten minutes. .tralized with acetic acid, and the mixture is then evaporated to dryness and flushed with n-hexane.

The excess sodium methoxide is neu- 3&5

Approximately 0.5 g. of 17a,20,20,2l-bis-methylenedioxy-l1,8-hydroxy-6,16a-dimethyl-4,6-pregnadiene-3 one is dissolved in 25 cc. of benzene, and about 5 cc. of benzene is removed from the solution by distillation at normal pressure. The resulting solution is cooled to room temperature, and to the solution is added about 0.75 cc. of freshly-distilled ethyl formate. The air in the system is replaced with nitrogen, and about mg. of sodium hydride (as a 57% dispersion in mineral oil) is added. The mixture is stirred under nitrogen at room temperature for a period of about three hours. About 15 cc. of a saturated aqueous solution sodium dihydrogen phosphate is added to the reaction mixture and the resulting aqueous mixture is extracted with ether. The benzene solution and ether extracts are combined, extracted with 2 N aqueous sodium hydroxide, and the aqueous sodium hydroxide extracts are acidified with sodium dihydrogen phosphate. The aqueous acidified solution is extracted with ether, the ether extract is evaporated to dryness, and the residual material is crystallized from ether to give 17a,20,20,2l-bismethylenedioxy 11,8 formyloxy 2 hydroxymethylene-6,16a-dimethyl-4,6-pregnadiene 3 one. Evaporation of the ether mother liquor,.followed by recrystallization of the residual material from ether gives substantially pure 17 a,20,20,2l-bismethylenedioxy-l 1 fl-hydroxy-Z-hydroxymethylene 6,16ot dimethyl-4,6-pregnadiene-3-one; M.P. 200-204 C.

Approximately 1.19 g. of 17a,20,20,2l-bismethylenedioxy-l 1 6-hydroxy-2-hydroxymethylene 6,16 dimethyl- 4,6-pregnadiene-3-one is dissolved in 25 cc. of ethanol. Three hundred mg. of phenyl hydrazine is added, and

the mixture is heated under reflux in a nitrogen atmosphere for one hour. About 25 cc. of water is added. The product is then extracted into 150 cc. of ether. The ether extracts are Washed with 2 N aqueous HQ, with saturated sodium bicarbonate, with water, with saturated aqueous sodium chloride solution, and then dried over anhydrous sodium sulfate. The washed and dried ether extracts are evaporated to dryness and the residual material is crystallized from ether to give 17a,20,20,Zl-bi$- methylenedioxy-6,l6oc-dimethyl 2'-phenyl [3,2-c1pyrazolo-4,6-pragnadiene-llfi-ol; M.P. 258262 C.

Approximately 430 mg. of 17a,20,20,2l-bismethylenedioxy-6,16a dimethyl 2'-phenyl [3,2-c]pyrazolo 4,6- pregnadiene-llfi-ol is heated on a steam bath under nitrogen with 40 cc. of a 60% aqueous solution of formic acid for a period of about 30 minutes. About 40 cc. of water is added to the reaction mixture, and the mixture is then extracted into about 200 cc. of chloroform. The chloroform solution is washed with water, with saturated aqueous sodium bicarbonate solution, again with water, and is then dried over anhydrous sodium sulfate. The Washed and dried chloroform solution is evaporated under vacuum and the residual product is dissolved in 60 cc. of absolute methanol, and 0.1 equivalent of sodium methoxide in methanol is added. The resulting mixture is stirred under nitrogen at room temperaturefor 15 minutes. The reaction mixture is acidified with acetic acid, and the solvent is evaporated under vacuum at room temperature. About 20 cc. of Water is added to the residual material, and the aqueous mixture is extracted with about 150cc. of ethyl acetate. The ethyl acetate solution is washed With saturated aqueous sodium bicarbonate solution, with water, and is then dried over anhydrous sodium sulfate and evaporated to dryness to give an amorphous solid. This amorphous solid is dried in a high vacuum, and is then dissolved in 4 cc. of pyridine. About 3 cc. of acetic anhydride is added. The resulting mixture is heated on .the steam bath for about 15 mintues and evaporated to dryness in vacuo. About 20 cc. of water is added. The

aqueous mixture is then extracted with about-150 cc. of

from ethyl acetate-benzene to give substantially pure 11fi,17a,21 trihydroxy 6,160; dimethyl 2 phenyl- [3,2-c]pyrazolo 4,6 pregnadiene 20 one 21-acetate; M.P. 225-226 C.

One hundred mg. of 11B,17a,21-trihydroxy-6,-l6u-dimethyl-2-phenyl-[3,2-c1pyrazolo 4,6 pregnadiene 20- one 21-acetate is dissolved in 10 cc. of absolute methanol, and 1.1 equivalent of sodium methoxide in methanol is added. The mixture is stirred at room temperature under nitrogen for 15 minutes. 'The product is acidified with acetic acid and is then evaporated at room temperature under vacuum. The residual material is dissolved in 100 cc. of chloroform, the chloroform solution is washed with water, with saturated aqueous sodium bicarbonate solution, again with water, and is then dried over anhydrous sodium sulfate. The washed and dried chloroform solution is evaporated to dryness to give 11,8,17a,21-trihydroxy-6,16a-dimethyl 2. phenyl [3,2-c]pyrazolo 4,6- pregnadiene-ZO-one.

Various changes and modifications may be made in carrying out the present invention without departing from the spirit and scope thereof. Insofar as these changes and modifications are within the purview of the annexed claims, they are to be considered as part of our invention.

We claim:

1. Sodium salt of 1113,17a,2l-trihydroxy-1,4-pregnadiene-3,20-dine 21-glucuronide.

2. Sodium salt of 16a-methyl-llfi,l7u,2l-trihydroxy-4- pregnene-3,20-di0ne 21-glucuronide.

3. Sodium salt of 16a-methyl-11fi,17a,21-trihydroxy- 1,4-pregnadiene-3,20 dione 21-g1ucuronide.

4. Sodium salt of 9a-flllOIO-16oc-m6thyl-11fi,17u.,21-t1'lhydroxy-1,4-pregnadiene-3,ZO-dione 2l-glucuronide.

5. Sodium salt of 9a-fluoro-16B-methyl-11B,17a,2l-trihydroxy-l,4-pregnadiene-3,20-dione 21-glucuronide.

. rated in vacuo, and the residual material is crystallized 6. Sodium salt of 6a-methyl-1 1B,17a,21-trihydroxy-1,4- pregnadiene-3,20-dione 21 glucuronide.

7. Sodium salt of a 6a-fluoro-11,B,l7u,21-trihydroxy- 1,4-pregnadiene-3,20-dione 21-glucuronide.

8. Sodium salt of 6,16a-dimethyl-11 8,17a,21-trihydroxy-4,6-pregnadiene-3,ZO-dione 21-glucur0nide.

9. A compound selected from the group consisting of 90a fluoro-l1p,16a,17a,21-tetrahydroxy-1,4-pregnadiene- 3,20-dione 2l-glucuronide, the 16,17-acetonide thereof, and pharmacologically acceptable salts of the foregoing.

10. Sodium salt of 6,16a-dimethyl-l1B,l7a,2l-trihydroxy [3,2-c]pyrazolo-4,6-pregnadiene-20-one 21-glucuronide.

11. Sodium salt of 6,l6oc-dimethyl-l1B,17a,21-trihydroXy-2-phenyl [3,2-c]pyrazolo-4,6-pregnadiene-20-one 21-glucuronide.

1'2. Methyl (11,13,170; dihydroxy-1,4-pregnadiene-3,20 dione-21-yl tri-O-acetyl-fi-D-glucopyranosid)-ur0nate.

13. Methyl(16a methyl 11,8,17u-dihydroxy-4-pregnene 3,20-dione-21-yl tri-O-acetyl-fi-D-glucopyranosid)- uronate.

l4. Methyl l6a-methyl-1 1fi,17a-dihydroxy-1,4-pregnadiene-3,20-dione-21-yl tri-O-acetyl-fi-D-glucopyranosid)- uronate.

15. Methyl (9u-fiu0ro-16a-methyl-1l,B,l 7a-dihydroxy- 1,4pregnadiene 3,20-dione-21-yl tri-O-acetyl-fl-D-glucopyranosid)-uronate.

16. Methyl ,(6,l6a-dimethyl-1lfl,l7a-dihydroxy[3,2-c] pyrazolo 4,6-pregnadiene-20-one-2l-yl tri-O-acetyl-fi-D- glucopyranosid)-uronate.

References Cited by the Examiner Artz et al.: Biochemistry of Glucuronic Acid (1950), pages -85, Academic Press Inc., New York.

. LEWIS GOTTS, Primary Examiner.

MORRIS LIEBMAN, Examiner. 

1. SODIUM SALT OF 11B,17A, 21-TRIHYDROXY-1.4-PREGNADIENE-3,20-DIONE 21-GLUCURONIDE. 